MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'genesis' source. Sat, 20 Mar 2010 15:47:09 +0100 http://www.medworm.com/index.php?rid=3345642&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20612 Transgenic mice are highly valuable tools for biological research as they allow cell type-specific expression of functionally instrumental genes. In this protocol, the generation of bacterial artificial chromosome (BAC) transgenic constructs is described. We give an overview of different transgenic inserts, such as fluorescent proteins (alone or in combination with Cre variants), diphtheria toxin receptor, lacZ, and light-activated ion channels. The most reliable and versatile approach to express these genes is by using BACs, which allow "highjacking" of the expression pattern of a gene without characterizing its transcriptional control elements. Here, we describe the necessary cloning techniques compared with conventional transgenesis. With the provided "toolbox" of already available tran... genesis journals http://www.medworm.com/rss/comments.php?id=3345642 Tue, 09 Mar 2010 00:00:00 +0100 3345642 http://www.medworm.com/index.php?rid=3321300&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20616 No abstract. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=3321300 Tue, 02 Mar 2010 00:00:00 +0100 3321300 http://www.medworm.com/index.php?rid=3321301&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20609 Two main classes of proteins, Polycomb group (PcG) and Trithorax group (TrxG), play a key role in the regulation of homeotic genes. These proteins act in multimeric complexes to remodel chromatin. A third class of proteins named Enhancers of Trithorax and Polycomb (ETP) modulates the activity of TrxG and PcG, but their role remains largely unknown. We previously identified an HMGB-like protein, DSP1 (Dorsal Switch Protein 1), which was classified as an ETP. Preliminary studies have revealed that DSP1 is involved in multimeric complexes. Here we identify a DEAD-box RNA helicase, Rm62, as partner of DSP1 in a 250-kDa complex. Coimmunoprecipitation assays performed on embryo extracts indicate that DSP1 and Rm62 are associated in 3- to 12-h embryos. Furthermore, DSP1 and Rm62 colocalize on pol... genesis journals http://www.medworm.com/rss/comments.php?id=3321301 Mon, 01 Mar 2010 00:00:00 +0100 3321301 http://www.medworm.com/index.php?rid=3317696&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20610 Formation of polyploid organisms by fertilization of unreduced gametes in meiotic mutants is believed to be a common phenomenon in species evolution. However, not well understood is how species in nature generally exist as haploid and diploid organisms in a long evolutionary time while polyploidization must have repeatedly occurred via meiotic mutations. Here, we show that the ploidy increased for two consecutive generations due to unreduced but viable gametes in the Arabidopsis cyclin a1;2-2 (also named tardy asynchronous meiosis-2) mutant, but the resultant octaploid plants produced progeny of either the same or reduced ploidy via genomic reductions during meiosis and pollen mitosis. Ploidy reductions through sexual reproduction were also observed in independently generated artificial oc... genesis journals http://www.medworm.com/rss/comments.php?id=3317696 Mon, 01 Mar 2010 00:00:00 +0100 3317696 http://www.medworm.com/index.php?rid=3297495&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20606 Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family of neurotrophic factors, has important functions in the peripheral and central nervous system of vertebrates. We have generated bacterial artificial chromosome (BAC) transgenic mice harboring 207 kb of the rat BDNF (rBDNF) locus containing the gene, 13 kb of genomic sequences upstream of BDNF exon I, and 144 kb downstream of protein encoding exon IX, in which protein coding region was replaced with the lacZ reporter gene. This BDNF-BAC drove transgene expression in the brain, heart, and lung, recapitulating endogenous BDNF expression to a larger extent than shorter rat BDNF transgenes employed previously. Moreover, kainic acid induced the expression of the transgenic BDNF mRNA in the cerebral cortex and hippocamp... genesis journals http://www.medworm.com/rss/comments.php?id=3297495 Tue, 23 Feb 2010 00:00:00 +0100 3297495 http://www.medworm.com/index.php?rid=3257786&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20607 Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder presenting with an elfin-like face, supravalvular aortic stenosis, a specific cognitive-behavioral profile, and infantile hypercalcemia. We encountered two WBS patients presenting with infantile spasms, which is extremely rare in WBS. Array comparative genomic hybridization (aCGH) and fluorescent in situ hybridization (FISH) analyses revealed atypical 5.7-Mb and 4.1-Mb deletions at 7q11.23 in the two patients, including the WBS critical region and expanding into the proximal side and the telomeric side, respectively. On the proximal side, AUTS2 and CALN1 may contribute to the phenotype. On the telomeric side, there are two candidate genes HIP1 and YWHAG. Because detailed information of them was unavailable, we investigated the... genesis journals http://www.medworm.com/rss/comments.php?id=3257786 Wed, 10 Feb 2010 00:00:00 +0100 3257786 http://www.medworm.com/index.php?rid=3257788&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20604 We have characterized a transgenic mouse line in which enhanced green fluorescent protein (EGFP) is expressed under the control of multimerized LEF-1 responsive elements. In embryos, EGFP was detected in known sites of Wnt activation, including the primitive streak, mesoderm, neural tube, somites, heart, limb buds, mammary placodes, and whisker follicles. In vitro cultured transgenic embryonic fibroblasts upregulated EGFP expression in response to activation of Wnt signaling by GSK3[beta] inhibition. Mammary tumor cell lines derived from female LEF-EGFP transgenic mice treated with the carcinogen 7, 12-dimethylbenz[a]anthracene (DMBA) also express EGFP. Thus, this transgenic line is useful for ex vivo and in vitro studies of Wnt signaling in development and cancer. genesis 00:1-12, 2010. Â... genesis journals http://www.medworm.com/rss/comments.php?id=3257788 Tue, 09 Feb 2010 00:00:00 +0100 3257788 http://www.medworm.com/index.php?rid=3257787&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20605 Cellular immortalization provides a way for expansion and subsequent molecular characterization of rare cell types. Ideally, immortalization can be achieved by the reversible expression of immortalizing proteins. Here, we describe the use of conditional immortalization based on a modified tetracycline-regulated system for the expression of SV40 large T-antigen in embryonic stem (ES) cells and mice. The modified system relies on a codon improved reverse tetracycline transactivator (irtTA) fused to the ligand-binding domain (LBD) of the androgen receptor (irtTA-ABD) or of a mutated glucocorticoid receptor (irtTA-GBD*). Induction of T-antigen is conferred only after addition of two ligands, one to activate the LBD (mibolerone for irtTA-ABD or dexamethasone for irtTA-GBD*) and one to activate ... genesis journals http://www.medworm.com/rss/comments.php?id=3257787 Tue, 09 Feb 2010 00:00:00 +0100 3257787 http://www.medworm.com/index.php?rid=3242734&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20601 Proteolytical cleavage of the [beta]-amyloid precursor protein (APP) generates [beta]-amyloid, which is deposited in the brains of patients suffering from Alzheimer's disease (AD). Despite the well-established key role of APP for AD pathogenesis, the physiological function of APP and its close homologues APLP1 and APLP2 remains poorly understood. Previously, we generated APP-/- mice that proved viable, whereas APP-/-APLP2-/- mice and triple knockouts died shortly after birth, likely due to deficits of neuromuscular synaptic transmission. Here, we generated conditional knockout alleles for both APP and APLP2 in which the promoter and exon1 were flanked by loxP sites. No differences in expression were detectable between wt and floxed alleles, whereas null alleles were obtained upon crossing ... genesis journals http://www.medworm.com/rss/comments.php?id=3242734 Fri, 05 Feb 2010 00:00:00 +0100 3242734 http://www.medworm.com/index.php?rid=3230451&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20603 Sall1 is expressed in the metanephric mesenchyme in the developing kidney, and mice deficient in Sall1 show kidney agenesis or dysgenesis. Sall1 is also expressed elsewhere, including in the limb buds, anus, heart, and central nervous system. Dominant-negative mutations of Sall1 in mice and humans lead to developmental defects in these organs. Here, we generated a mouse line expressing tamoxifen-inducible Cre recombinase (CreERT2) under the control of the endogenous Sall1 promoter. Upon tamoxifen treatment, these mice showed genomic recombination in the tissues where endogenous Sall1 is expressed. When CreERT2 mice were crossed with the floxed Sall1 allele, tamoxifen administration during gestation led to a significant decrease in Sall1 expression and small kidneys at birth, suggesting tha... genesis journals http://www.medworm.com/rss/comments.php?id=3230451 Tue, 02 Feb 2010 00:00:00 +0100 3230451 http://www.medworm.com/index.php?rid=3230452&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20602 The goal of this study was to determine the presence of osteoprogenitor cells in the peripheral blood. Experiments were conducted with a parabiosis model in which osteoblast specific transgenic mice (Col2.3GFP or hOC-GFP) were surgically joined with a transgenic mouse where herpes virus thymidine kinase gene is under the control of the collagen [alpha]1 promoter (Col2.3[Delta]TK). This method permits conditional ablation of osteoblasts by ganciclovir (GCV) treatment. In parabionts treated with GCV for 15 days or 1.5-2 months, GFP (hOC-GFP or Col2.3GFP) expression was not detected in histological preparations or in marrow stromal cell cultures from the Col2.3[Delta]TK parabiont. Finally, Col2.3GFP/Col2.3[Delta]TK pairs were treated with GCV for 15 days and allowed to recover from GCV for 3 ... genesis journals http://www.medworm.com/rss/comments.php?id=3230452 Mon, 01 Feb 2010 00:00:00 +0100 3230452 http://www.medworm.com/index.php?rid=3211094&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20599 Notch signaling is essential for embryonic vascular development in mammals and other vertebrates. Here we show that mouse embryos with conditional activation of the Notch1 gene in endothelial cells (Notch1 gain of function embryos) exhibit defects in vascular remodeling increased diameter of the dorsal aortae, and form arteriovenous malformations. Conversely, embryos with either constitutive or endothelial cell-specific Notch1 gene deletion also have vascular defects, but exhibit decreased diameter of the dorsal aortae and form arteriovenous malformations distinctly different from the Notch1 gain of function mutants. Surprisingly, embryos homozygous for mutations of the ephrinB/EphB pathway genes Efnb2 and Ephb4 exhibit vascular defects and arteriovenous malformations that phenocopy the No... genesis journals http://www.medworm.com/rss/comments.php?id=3211094 Wed, 27 Jan 2010 00:00:00 +0100 3211094 http://www.medworm.com/index.php?rid=3196657&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20600 In this report, we describe a genetic interaction between nanos and Enhancer of nos, an allele of the previously uncharacterized locus CG4699. We find that E(nos) protein is required for normal accumulation of Nanos in the ovary and thus for maintenance of the germ line. The mechanism by which E(nos)/CG4699 protein acts is not clear, although it has been found in a complex with Mof acetylase. Consistent with the finding that E(nos) interacts with Mof, we observe that nanos and mof also interact genetically to maintain normal oogenesis. genesis 00:1-10, 2010. © 2010 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=3196657 Fri, 22 Jan 2010 00:00:00 +0100 3196657 http://www.medworm.com/index.php?rid=3196661&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20595 Genome engineering strategies employing site-specific recombinases (SSRs) have become invaluable to the study of gene function in model organisms. One such SSR, the integrase encoded by the Streptomyces bacteriophage phiC31, promotes recombination between heterotypic attP and attB sites. In the present study I have examined the feasibility of the use of phiC31 integrase for intramolecular recombination strategies in zebrafish embryos. I report here that (1) phiC31 integrase is functional in zebrafish cells, (2) phiC31 integrase can excise a transgene cassette flanked by an attB and an attP site, analogous to a common use of the Cre/lox SSR system, (3) phiC31 integrase functions in the zebrafish germline, and (4) a phiC31 integrase-estrogen receptor hormone-binding domain variant fusion pro... genesis journals http://www.medworm.com/rss/comments.php?id=3196661 Thu, 21 Jan 2010 00:00:00 +0100 3196661 http://www.medworm.com/index.php?rid=3196660&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20594 After the successful completion of the human genome project (HGP), biological research in the postgenome era urgently needs an efficient approach for functional analysis of genes. Utilization of knockout mouse models has been powerful for elucidating the function of genes as well as finding new therapeutic interventions for human diseases. Gene trapping and gene targeting are two independent techniques for making knockout mice from embryonic stem (ES) cells. Gene trapping is high-throughput, random, and sequence-tagged while gene targeting enables the knockout of specific genes. It has been about 20 years since the first gene targeting and gene trapping mice were generated. In recent years, new tools have emerged for both gene targeting and gene trapping, and organizations have been formed... genesis journals http://www.medworm.com/rss/comments.php?id=3196660 Thu, 21 Jan 2010 00:00:00 +0100 3196660 http://www.medworm.com/index.php?rid=3196659&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20596 The T-box transcription factor Tbx2 plays important roles in patterning and development, and has been implicated in cell-cycle regulation and cancer. Conventional disruption of Tbx2 results in abnormalities of the heart, limbs, eye and other structures, and early fetal lethality. To gain insight into the role of Tbx2 in different tissues and at different stages of development, we have generated a conditional null allele of Tbx2 by flanking Exon 2 with loxP sites (Tbx2fl2). Homozygous Tbx2fl2 mice are viable and fertile, indicating that the Tbx2fl2 allele is a fully functional Tbx2 allele. Cre-mediated recombination, using a ubiquitously active CMV-Cre line, results in deletion of Exon 2 and loss of protein expression. Embryos homozygous for the recombined allele (Tbx2[Delta]2) show the sam... genesis journals http://www.medworm.com/rss/comments.php?id=3196659 Thu, 21 Jan 2010 00:00:00 +0100 3196659 http://www.medworm.com/index.php?rid=3196658&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20598 Transgenic mice are vital tools in both basic and applied research. Unfortunately, the transgenesis process as well as many other assisted reproductive techniques involving embryo transfer rely on vasectomized males to induce pseudopregnancy in surrogate mothers. Vasectomy is a surgical procedure associated with moderate pain and must be carried out under full anaesthesia by qualified personnel. Eliminating the need for vasectomy would be beneficial from the economic and animal welfare point of view. Our aim was to develop a transgene-based alternative to the surgical vasectomy procedure. We generated several transgenic mouse lines expressing a Protamine-1 (Prm1) EGFP fusion protein under the transcriptional and translational regulatory control of Prm1. Male mice from lines showing moderat... genesis journals http://www.medworm.com/rss/comments.php?id=3196658 Thu, 21 Jan 2010 00:00:00 +0100 3196658 http://www.medworm.com/index.php?rid=3141506&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20590 CLIC1 belongs to a family of highly conserved and widely expressed intracellular chloride ion channel proteins existing in both soluble and membrane integrated forms. To study the physiological and biological role of CLIC1 in vivo, we undertook conditional gene targeting to engineer Clic1 gene knock-out mice. This represents creation of the first gene knock-out of a vertebrate CLIC protein family member. We first generated a Clic1 Knock-in (Clic1FN) allele, followed by Clic1 knock-out (Clic1-/-) mice by crossing Clic1FN allele with TNAP-cre mice, resulting in germline gene deletion through Cre-mediated recombination. Mice heterozygous or homozygous for these alleles are viable and fertile and appear normal. However, Clic1-/- mice show a mild platelet dysfunction characterized by prolonged ... genesis journals http://www.medworm.com/rss/comments.php?id=3141506 Tue, 05 Jan 2010 00:00:00 +0100 3141506 http://www.medworm.com/index.php?rid=3113987&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20586 Considering the regulatory complexities of progesterone receptor (PR) action throughout the female reproductive axis and mammary gland, we generated a mouse model that enables conditional ablation of PR function in a spatiotemporal specific manner. Exon 2 of the murine PR gene was floxed to generate a conditional PR allele (PRflox) in mice. Crossing the PRflox/flox mouse with the ZP3-cre transgenic demonstrated that the PRflox allele recombines to a PR null allele (PRd). Mice homozygous for the recombined null PR allele (PRd/d) exhibit uterine, ovarian, and mammary gland defects that phenocopy those of our previously described PR knockout (PRKO) model. Therefore, this conditional mouse model for PR ablation represents an invaluable resource with which to further define in a developmental a... genesis journals http://www.medworm.com/rss/comments.php?id=3113987 Tue, 22 Dec 2009 00:00:00 +0100 3113987 http://www.medworm.com/index.php?rid=3110370&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20589 Loss of Rb1 tumor suppressor gene function is involved in the genesis of most human cancers. Novel therapies targeting Rb1 have been slow to develop because of our incomplete understanding of its molecular mechanisms of action. Rb1 protein (pRb) binds a host of cellular genes and proteins, and these molecular interactions mediate its various functions. Given the potential complexity of these molecular interactions and the lack of established methods for pRb purification, it has been difficult to systematically identify gene and protein interactions relevant to tumor suppression in different tissues in vivo. To address this limitation, we have generated a dual affinity tagged Rb1 allele in the mouse. The tagged allele functions as wild type and the encoded protein can be purified by tandem ... genesis journals http://www.medworm.com/rss/comments.php?id=3110370 Tue, 22 Dec 2009 00:00:00 +0100 3110370 http://www.medworm.com/index.php?rid=3110372&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20583 Lens formation in mouse is critically dependent on proper development of the retinal neuroectoderm that is located close beneath the head surface ectoderm. Signaling from the prospective retina triggers lens-specific gene expression in the surface-ectoderm. Supression of canonical Wnt/[beta]-catenin signaling in the surface ectoderm is one of the prerequisites for lens development because, as we show here, ectopic Wnt activation in the retina and lens abrogates lens formation. Wnt inhibiton is mediated by signals coming from the retina but its exact mechanism is unknown. We show that Pax6 directly controls expression of several Wnt inhibitors such as Sfrp1, Sfrp2, and Dkk1 in the presumptive lens. In accordance, absence of Pax6 function leads to aberrant canonical Wnt activity in the presu... genesis journals http://www.medworm.com/rss/comments.php?id=3110372 Mon, 21 Dec 2009 00:00:00 +0100 3110372 http://www.medworm.com/index.php?rid=3110371&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20588 We generated transgenic mouse line C57BL/6-Tg(Hspa2-cre)1Eddy/J (Hspa2-cre), which expresses cre-recombinase under the control of a 907-bp fragment of the heat shock protein 2 (Hspa2) gene promoter. Transgene expression was determined using Gt(ROSA)26Sortm1Sor/J (ROSA26) and Tg(CAG-Bgeo/GFP)21Lbe/J (Z/EG) reporter strains and RT-PCR and immunohistochemistry assays. Hspa2-cre expression mimicked the spermatogenic cell-specific expression of endogenous HSPA2 within the testis, being first observed in leptotene/zygotene spermatocytes. Expression of the transgene also was detected at restricted sites in the brain, as occurs for endogenous HSPA2. Although the results of mating the Hspa2-cre mice to mice with a floxed Cdc2a allele indicated that some expression of the transgene occurs during emb... genesis journals http://www.medworm.com/rss/comments.php?id=3110371 Mon, 21 Dec 2009 00:00:00 +0100 3110371 http://www.medworm.com/index.php?rid=3095609&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20579 Bone morphogenetic proteins (BMPs) have important functions during epithelial development. In Drosophila, extracellular Short gastrulation (Sog) limits the action of the BMP family member Decapentaplegic (Dpp). We have shown that Integrin receptors regulate Sog activity and distribution during pupal wing development to direct placement of wing veins. Here, we show that Integrins perform a similar function in the follicular epithelium, impacting Dpp function during oogenesis and embryonic development. As reported for the wing, this effect is specific to mew, which codes for [alpha]PS1 integrin. Sog is subject to cleavage by metalloproteases, generating fragments with different properties. We also show that Integrins regulate the distribution of C- and N-terminal Sog fragments in both epithe... genesis journals http://www.medworm.com/rss/comments.php?id=3095609 Thu, 17 Dec 2009 00:00:00 +0100 3095609 http://www.medworm.com/index.php?rid=3091712&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20587 Ephrins and Eph receptor tyrosine kinases are cell-surface molecules that serve a multitude of functions in cell-cell communication in development, physiology, and disease. EphA4 is a promiscuous member of the EphA subclass of Eph receptors and can bind to both EphrinAs and EphrinBs. In addition to its well-established roles in guiding the development of neuronal connectivity, EphA4 has been implicated for a role in synaptic plasticity, vascular formation, axon regeneration, and central nervous system repair following injury. However, the study of its role in the adult stage has been hampered by confounding developmental defects in EphA4 germline mutants. Here, we report the generation and molecular characterization of an EphA4 conditional allele along with a novel null allele with a knock... genesis journals http://www.medworm.com/rss/comments.php?id=3091712 Wed, 16 Dec 2009 00:00:00 +0100 3091712 http://www.medworm.com/index.php?rid=3091718&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20573 We developed a conditional and inducible gene knockout methodology that allows effective gene deletion in mouse cardiomyocytes. This transgenic mouse line was generated by coinjection of two transgenes, a "reverse" tetracycline-controlled transactivator (rtTA) directed by a rat cardiac troponin T (Tnnt2) promoter and a Cre recombinase driven by a tetracycline-responsive promoter (TetO). Here, Tnnt2-rtTA activated TetO-Cre expression takes place in cardiomyocytes following doxycycline treatment. Using two different mouse Cre reporter lines, we demonstrated that expression of Cre recombinase was specifically and robustly induced in the cardiomyocytes of embryonic or adult hearts following doxycycline induction, thus, allowing cardiomyocyte-specific gene disruption and lineage tracing. We als... genesis journals http://www.medworm.com/rss/comments.php?id=3091718 Tue, 15 Dec 2009 00:00:00 +0100 3091718 http://www.medworm.com/index.php?rid=3091717&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20578 We report the detection of a physical interaction between Dyrk1b and Wdr68. We also found perturbations of nodal signaling in dyrk1b antisense morpholino knockdown (dyrk1b-MO) animals. Specifically, we found reduced expression of lft1 and lft2 (lft1/2) during gastrulation and a near complete loss of the later asymmetric lft1/2 expression domains. Although wdr68-MO animals did not display lft1/2 expression defects during gastrulation, they displayed a near complete loss of the later asymmetric lft1/2 expression domains. While expression of ndr1 was not substantially effected during gastrulation, ndr2 expression was moderately reduced in dyrk1b-MO animals. Analysis of additional downstream components of the nodal signaling pathway in dyrk1b-MO animals revealed modestly expanded expression of... genesis journals http://www.medworm.com/rss/comments.php?id=3091717 Tue, 15 Dec 2009 00:00:00 +0100 3091717 http://www.medworm.com/index.php?rid=3091716&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20581 Analysis of the retinal defects of a CK2 phosphomimetic variant of E(spl)M8 (M8S159D) and the truncated protein M8* encoded by the E(spl)D allele, suggest that the nonphosphorylated CtD "autoinhibits" repression. We have investigated this model by testing for inhibition (in "trans") by the CtD fragment in its nonphosphorylated (M8-CtD) and phosphomimetic (M8SD-CtD) states. In N+ flies, ectopic M8-CtD compromises lateral inhibition, i.e., elicits supernumerary bristles as with loss of N signaling. This antimorphic activity of M8-CtD strongly rescues the reduced eye and/or bristle loss phenotypes that are elicited by ectopic M8SD or wild type M8. Additionally, the severely reduced eye of Nspl/Y; E(spl)D/+ flies is also rescued by M8-CtD. Rescue is specific to the time and place, the morphoge... genesis journals http://www.medworm.com/rss/comments.php?id=3091716 Tue, 15 Dec 2009 00:00:00 +0100 3091716 http://www.medworm.com/index.php?rid=3091715&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20584 TDP-43 is a highly conserved and ubiquitously expressed nuclear protein. It has been implicated in the regulation of transcription, alternative splicing, translation, and neuronal plasticity. TDP-43 has also been shown to be a disease signature protein associated with several neurodegenerative diseases including amyotrophic lateral sclerosis. However, the correlation of the physiological functions of TDP-43 with these diseases remains unknown. We have used the gene targeting approach to disrupt the expression of TDP-43 in mouse. Loss of the TDP-43 expression results in peri-implantation lethality of mice between embryonic days (E) 3.5 and 6.5. Blastocysts of the homozygous Tardbp null mutants are morphologically normal, but exhibit defective outgrowth of the inner cell mass in vitro. Our d... genesis journals http://www.medworm.com/rss/comments.php?id=3091715 Tue, 15 Dec 2009 00:00:00 +0100 3091715 http://www.medworm.com/index.php?rid=3091714&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20585 The Z/EG transgenic mouse line, produced by Novak et al., displays tissue-specific EGFP expression after Cre-mediated recombination. The autofluorescence of EGFP allows the visualization of cells of interest displaying Cre recombination. The initial construct was designed such that cells without Cre recombination express the [beta]-galactosidase marker, facilitating counterselection. We used inverse PCR to identify the site of integration of the Z/EG transgene, to improve the efficiency of homozygous Z/EG mouse production. Recombined cells produced large amounts of EGFP protein, resulting in higher levels of fluorescence and therefore greater contrast with nonrecombined cells. We mapped the transgene to the G1 region of chromosome 5. This random insertion was found to have occurred 230-bp ... genesis journals http://www.medworm.com/rss/comments.php?id=3091714 Tue, 15 Dec 2009 00:00:00 +0100 3091714 http://www.medworm.com/index.php?rid=3091713&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20592 No abstract. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=3091713 Tue, 15 Dec 2009 00:00:00 +0100 3091713 http://www.medworm.com/index.php?rid=2946157&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20577 Accelerated vascular calcification occurs in several human diseases including diabetes and chronic kidney disease (CKD). In patients with CKD, vascular calcification is highly correlated with elevated serum phosphate levels. In vitro, elevated concentrations of phosphate induced vascular smooth muscle cell matrix mineralization, and the inorganic phosphate transporter-1 (PiT-1), was shown to be required. To determine the in vivo role of PiT-1, mouse conditional and null alleles were generated. Here we show that the conditional allele, PiT-1flox, which has loxP sites flanking exons 3 and 4, is homozygous viable. Cre-mediated recombination resulted in a null allele that is homozygous lethal. Examination of early embryonic development revealed that the PiT-1[Delta]e3,4/[Delta]e3,4 embryos dis... genesis journals http://www.medworm.com/rss/comments.php?id=2946157 Sat, 31 Oct 2009 00:00:00 +0100 2946157 http://www.medworm.com/index.php?rid=2946163&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20571 In Drosophila, the Gal4-UAS system is used to drive ectopic gene expression in a tissue-specific manner. In this system, transgenic flies expressing tissue specific Gal4 are crossed to a line in which the gene to be expressed is under the control of a Gal4-responsive UAS sequence. The resulting progeny express the gene of interest in the pattern of the particular Gal4 line. Since a given UAS-transgene can be driven by any Gal4 line, this system is predominantly limited by available Gal4 lines. Here we report the characterization of a novel line, DE-Gal4, which in the eye is expressed in the dorsal compartment for the majority of development. Furthermore, we use functional tests to show that the DE-Gal4 line is a useful tool with which to manipulate gene expression in half of the developing... genesis journals http://www.medworm.com/rss/comments.php?id=2946163 Fri, 30 Oct 2009 00:00:00 +0100 2946163 http://www.medworm.com/index.php?rid=2946162&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20567 Chondrichthyans (cartilaginous fishes) represent one of the two lineages of gnathostomes, the other being the osteicthyans (bony fishes). Classical studies on chondrichthyan embryology have strongly impacted our views of vertebrate body plan evolution, while recent studies highlight oviparous chondrichthyans as emerging vertebrate model systems that are amenable to experimental embryological manipulation. Here, we review three particular areas of interest in the field of chondrichthyan developmental biology - gastrulation, neural development, and appendage patterning - and we discuss recentfindings within a broader chondrichthyan-osteichthyan comparative framework. In some cases, comparative studies of chondrichthyan and osteichthyan development reveal conserved patterns of gene expression... genesis journals http://www.medworm.com/rss/comments.php?id=2946162 Fri, 30 Oct 2009 00:00:00 +0100 2946162 http://www.medworm.com/index.php?rid=2946161&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20572 This reporter transgene consisting of a humanized renilla Green Fluorescent Protein (hrGFP) sequence controlled by the Upstream Activating Sequences for the Gal4 gene (UASG) was inserted into the modified ER[alpha] BAC clone. Expression of Gal4-ER[alpha] and hrGFP reliably recapitulates endogenous ER[alpha] expression and activity in the estrogen target tissues in response to estrogen stimulation. Therefore, the ERAAI mouse represents a novel animal model to investigate dynamic ER[alpha] activity in vivo. genesis 00:1-10, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2946161 Fri, 30 Oct 2009 00:00:00 +0100 2946161 http://www.medworm.com/index.php?rid=2946160&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20574 The MHC class I allochimeric protein containing donor-type epitopes on recipient-type heavy chains induces indefinite survival of heterotopic cardiac allografts in rats. We analyzed gene expression profile of heart allograft tissue. Mutated peptide [[alpha]1h1/u]-RT1.Aa that contains donor-type (Wistar Furth, WF; RT1u) immunogenic epitopes displayed on recipient-type (ACI, RT1a) was delivered into ACI recipients of WF hearts at the time of transplantation in addition to a 3 days course of oral cyclosporine. Microarray analysis was performed using Affymetrix Rat 230 2.0 Microarray. Allochimeric molecule treatment caused upregulation of genes involved in structural integrity of heart muscle, downregulation of IL-1[beta] a key modulator of the immune response, and downregulation of partitioni... genesis journals http://www.medworm.com/rss/comments.php?id=2946160 Fri, 30 Oct 2009 00:00:00 +0100 2946160 http://www.medworm.com/index.php?rid=2946159&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20575 Textpresso Site Specific Recombinases () is a text-mining web server for searching a database of more than 9,000 full-text publications. The papers and abstracts in this database represent a wide range of topics related to site-specific recombinase (SSR) research tools. Included in the database are most of the papers that report the characterization or use of mouse strains that express Cre recombinase as well as papers that describe or analyze mouse lines that carry conditional (floxed) alleles or SSR-activated transgenes/knockins. The database also includes reports describing SSR-based cloning methods such as the Gateway or the Creator systems, papers reporting the development or use of SSR-based tools in systems such as Drosophila, bacteria, parasites, stem cells, yeast, plants, zebrafis... genesis journals http://www.medworm.com/rss/comments.php?id=2946159 Fri, 30 Oct 2009 00:00:00 +0100 2946159 http://www.medworm.com/index.php?rid=2946158&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20576 Recently we have shown that BhSGAMP-1 is a developmentally regulated reiterated gene that encodes an antimicrobial peptide (AMP) and is expressed exclusively in the salivary glands, at the end of the larval stage. We show, for the first time, that a gene for an AMP is directly activated by 20-OH ecdysone. This control probably involves the participation of short-lived repressor(s). We also found that the promoter of BhSGAMP-1 is not equipped with elements that respond to infection, provoked by the injection of microorganisms, in the salivary glands or in the fat body. We produced polyclonal antibodies against the synthetic peptide and found that the BhSGAMP-1 peptide is secreted in the saliva. The BhSGAMP-1 gene was also activated during the third larval molt. These facts confirm our hypot... genesis journals http://www.medworm.com/rss/comments.php?id=2946158 Fri, 30 Oct 2009 00:00:00 +0100 2946158 http://www.medworm.com/index.php?rid=2893466&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20569 Promoter selection is of utmost importance for the study of in vivo gene function using transgenic models. In the present study, we have analyzed the expression of the GFP marker under the control of the composite CAG promoter in the lymphoid compartment of several transgenic mouse strains. Despite the ability of the CAG promoter to drive gene expression in almost all tissues examined to date, its activity appears to be developmentally regulated within the T lymphocyte cell lineage. In particular, CD4 and CD8-expressing, thymic immature T cells displayed lower levels of the GFP marker when compared with both bone marrow precursors and mature circulating T cells, suggesting a transient downregulation of CAG activity during T cell development. Alternative promoters may therefore be preferred... genesis journals http://www.medworm.com/rss/comments.php?id=2893466 Wed, 14 Oct 2009 23:00:00 +0100 2893466 http://www.medworm.com/index.php?rid=2893475&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20582 HeLa cells transfected with plasmids expressing dual fluorescent protein reporters with cherry, green, and cyan fluorscent proteins. Histone 2B fusions localize fluorescent proteins to the nucleus, while fluorescent proteins fused to glycosylphosphatidylinositol anchor signal sequence localizes to plasma membranes. These dual fluorescent reporters label nuclear and plasma membranes of live cells to follow their behaviors in developing embryos. See the paper by Stewart et al. in this issue. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2893475 Tue, 13 Oct 2009 23:00:00 +0100 2893475 http://www.medworm.com/index.php?rid=2893474&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20558 We present mouse lines expressing an inducible Cre protein (MerCreMer) under the control of regulatory elements of the probasin gene on a BAC. These mouse lines show high organ specificity, high transgene expression in anterior, dorsal and lateral prostate lobes, no background Cre recombination using a reporter strain and adjustable amounts of Cre-induced recombination upon tamoxifen induction. Together with two recently reported transgenic lines expressing the Cre-ERT2 protein from small prostate-specific promoters, these mouse lines will be useful in research focused on prostate-specific disorders such as benign hyperplasia or cancer. genesis 00:1-8, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2893474 Tue, 13 Oct 2009 23:00:00 +0100 2893474 http://www.medworm.com/index.php?rid=2893473&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20559 Utilizing a recently identified Sox10 distal enhancer directing Cre expression, we report S4F:Cre, a transgenic mouse line capable of inducing recombination in oligodendroglia and all examined neural crest derived tissues. Assayed using R26R:LacZ reporter mice expression was detected in neural crest derived tissues including the forming facial skeleton, dorsal root ganglia, sympathetic ganglia, enteric nervous system, aortae, and melanoblasts, consistent with Sox10 expression. LacZ reporter expression was also detected in non-neural crest derived tissues including the oligodendrocytes and the ventral neural tube. This line provides appreciable differences in Cre expression pattern from other transgenic mouse lines that mark neural crest populations, including additional populations defined... genesis journals http://www.medworm.com/rss/comments.php?id=2893473 Tue, 13 Oct 2009 23:00:00 +0100 2893473 http://www.medworm.com/index.php?rid=2893472&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20561 Based on environmental challenges or altered genetic composition, Drosophila larvae can produce up to three types of blood cells that express genetic programs essential for their distinct functions. Using transcriptional enhancers for genes expressed exclusively in plasmatocytes, crystal cells, or lamellocytes, several new hemocyte-specific enhancer-reporter transgenes were generated to facilitate the analysis of Drosophila hematopoiesis. This approach took advantage of fluorescent variants of insulated P-element reporter vectors for multilabeling cell analyses; two additional color variants were generated in these studies. These vectors were successfully used to produce transgenic fly lines that label specific hemocyte lineages with separate colors. Combining three transgene reporters all... genesis journals http://www.medworm.com/rss/comments.php?id=2893472 Tue, 13 Oct 2009 23:00:00 +0100 2893472 http://www.medworm.com/index.php?rid=2893471&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20562 During mouse embryonic development, the T-box transcription factor Eomes/Tbr2 is expressed in highly dynamic patterns in various progenitor cell types. Those include the undifferentiated cells of the trophectoderm, ingressing nascent mesoderm at the primitive streak, and intermediate progenitor cells of the developing cerebral cortex. We generated an EomesGFP- targeted allele to follow the highly dynamic patterns of Eomes expression and to allow for the identification of novel expression domains. We show that our novel allele recapitulates endogenous gene expression at known sites of expression and confirm our results by anti-Eomes immunofluorescent staining. Using this novel allele we were able to identify previously undocumented domains of Eomes expression within the visceral endoderm an... genesis journals http://www.medworm.com/rss/comments.php?id=2893471 Tue, 13 Oct 2009 23:00:00 +0100 2893471 http://www.medworm.com/index.php?rid=2893470&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20563 Transgenic and knockout studies have advanced our understanding of the genetic control of embryonic development over the past decades. However, interpretation of the phenotype of mutant mice is potentially complicated, since the commonly used knockout approach modifies both the fetal and placental genome. To circumvent this problem, we previously developed a placenta-specific gene manipulation system by lentiviral vector transduction of embryos at the blastocyst stage. In the present study, by combination with the Cre/LoxP system, we successfully demonstrate placenta-specific gene activation and inactivation in EGFP reporter mice and Ets2 floxed mice, respectively. Transient expression using integrase-defective lentiviral (IDLV) vectors diminished the toxic effect of Cre expression and sol... genesis journals http://www.medworm.com/rss/comments.php?id=2893470 Tue, 13 Oct 2009 23:00:00 +0100 2893470 http://www.medworm.com/index.php?rid=2893469&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20564 In conclusion, we have generated a mouse model in which genes implicated in cartilage degenerative diseases can be inactivated in a spatial and temporal fashion in postnatal and adult mice. genesis 00:1-10, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2893469 Tue, 13 Oct 2009 23:00:00 +0100 2893469 http://www.medworm.com/index.php?rid=2893468&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20566 The Wnt family member Wnt4 is critical for the development of several organs, including the kidney, gonad, and adrenal, mammary, and pituitary glands. To study its potential postnatal functions, we generated a floxed Wnt4 allele. A single loxP site was targeted to the second intron, while a loxP-Neo-loxP cassette was placed 3[prime] from Exon 5. The floxed Neo cassette was subsequently removed by crossing with MeuCre40 transgenic mice. The Wnt4 gene was specifically inactivated with CAGCre and another Wnt4 allele, Wnt4EGFPCre, in which the Cre is driven by the endogenous Wnt4 promoter. Deletion of Wnt4 gene function with CAGCre impaired kidney development, as is the case with the conventional knockout. Similarly, the Wnt4EGFPCre-mediated inactivation of Wnt4 function considerably reduced t... genesis journals http://www.medworm.com/rss/comments.php?id=2893468 Tue, 13 Oct 2009 23:00:00 +0100 2893468 http://www.medworm.com/index.php?rid=2893467&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20568 The albCre transgene, having Cre recombinase driven by the serum albumin (alb) gene promoter, is commonly used to generate adult mice having reliable hepatocyte-specific recombination of loxP-flanked ("floxed") alleles. Based on previous studies, it has been unclear whether albCre transgenes are also reliable in fetal and juvenile mice. Perinatal liver undergoes a dynamic transition from being predominantly hematopoietic to predominantly hepatic. We evaluated Cre activity during this transition in albCre mice using a sensitive two-color fluorescent reporter system. From fetal through adult stages, in situ patterns of Cre-dependent recombination of the reporter closely matched expression of endogenous Alb mRNA or protein, indicating most or all hepatocytes, including those in fetal and juve... genesis journals http://www.medworm.com/rss/comments.php?id=2893467 Tue, 13 Oct 2009 23:00:00 +0100 2893467 http://www.medworm.com/index.php?rid=2870956&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20565 Fluorescent proteins (FPs) are useful tools for visualizing live cells and their behaviors. Protein domains that mediate subcellular localization have been fused to FPs to highlight cellular structures. FPs fused with histone H2B incorporate into chromatin allowing visualization of nuclear events. FPs fused to a glycosylphosphatidylinositol anchor signal sequence label the plasma membrane, highlighting cellular shape. Thus, a reporter gene containing both types of FP fusions would allow for effective monitoring of cell shape, movement, mitotic stage, apoptosis, and other cellular activities. Here, we report a binary color-coding system using four differently colored FP reporters that generates 16 distinct color codes to label the nuclei and plasma membranes of live cells in culture and in ... genesis journals http://www.medworm.com/rss/comments.php?id=2870956 Wed, 07 Oct 2009 23:00:00 +0100 2870956 http://www.medworm.com/index.php?rid=2690583&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20557 The very limited ability to regenerate axons after injury in the mature mammalian central nervous system (CNS) has been partly attributed to the growth restrictive nature of CNS myelin. Oligodendrocyte myelin glycoprotein (OMgp) was identified as a major myelin-derived inhibitor of axon growth. However, its role in axon regeneration in vivo is poorly understood. Here we describe the generation and molecular characterization of an OMgp allelic series. With a single gene targeting event and Cre/FLP mediated recombination, we generated an OMgp null allele with a LacZ reporter, one without a reporter gene, and an OMgp conditional allele. This allelic series will aid in the study of OMgp in adult CNS axon regeneration using mouse models of spinal cord injury. The conditional allele will overcom... genesis journals http://www.medworm.com/rss/comments.php?id=2690583 Tue, 11 Aug 2009 23:00:00 +0100 2690583 http://www.medworm.com/index.php?rid=2690584&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20555 Spectrins are major proteins in the cytoskeletal network of most cells. In Drosophila, [beta]Heavy-Spectrin encoded by the karst gene functions together with Crb during photoreceptor morphogenesis. However, the roles of two other Spectrins ([alpha]- and [beta]-Spectrins) in developing photoreceptor cells have not been studied. Here, we analyzed the effects of spectrin mutations on developing eyes to determine their roles in photoreceptor morphogenesis. We found that the Spectrins are dispensable for retinal differentiation in eye imaginal discs during larval stage. However, photoreceptors deficient in [alpha]- or [beta]-Spectrin display dramatic apical membrane expansions including Crb and show morphogenesis defects during pupal eye development, suggesting that [alpha]- and [beta]-Spectrin... genesis journals http://www.medworm.com/rss/comments.php?id=2690584 Mon, 10 Aug 2009 23:00:00 +0100 2690584 http://www.medworm.com/index.php?rid=2656415&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20554 In this report, we describe a novel light- activatable knockdown reagent called PhotoMorphTM. PhotoMorphs can be generated from existing morpholinos by hybridization with a complementary caging strand containing a photocleavable linkage. The caging strand neutralizes the morpholino activity until irradiation of the PhotoMorph with UV light releases the morpholino. We generated PhotoMorphs to target genes encoding enhanced green fluorescent protein, No tail, and E-cadherin to illustrate the utility of this approach. Temporal control of gene expression with PhotoMorphs permitted us to circumvent the early lethal phenotype of E-cadherin knockdown. A splice-blocking PhotoMorph directed to the rheb gene showed light-dependent gene knockdown up to 72 hpf. PhotoMorphs thus offer a new class of la... genesis journals http://www.medworm.com/rss/comments.php?id=2656415 Thu, 30 Jul 2009 23:00:00 +0100 2656415 http://www.medworm.com/index.php?rid=2619619&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20553 Periphilin is involved in multiple processes in vivo. To explore its physiological role from an organismic perspective, we generated mice with a gene trap insertion in the periphilin-1 gene. Based on [beta]-gal reporter activity, a widespread periphilin expression was evident, especially in the developing somites and limbs, the embryonic nervous system, and the adult brain. In accordance with this broad expression, homozygous deficiency of periphilin was lethal in early embryogenesis. Mice with a heterozygous deficiency did not show any abnormalities of brain morphology and function, neither histologically nor regarding the transcriptome. Interestingly, the reduction of the periphilin-1 gene dosage was compensated by an increased expression of the remaining wild-type allele in the brain. T... genesis journals http://www.medworm.com/rss/comments.php?id=2619619 Mon, 20 Jul 2009 23:00:00 +0100 2619619 http://www.medworm.com/index.php?rid=2619624&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20544 Cortactin is an F-actin binding protein that has been suggested to play key roles in various cellular functions. Here, we generated mice carrying floxed alleles of the cortactin (Cttn) gene (Cttnflox/flox mice). Expression of Cre recombinase in mouse embryonic fibroblasts (MEFs) isolated from Cttnflox/flox embryos depleted cortactin within days, without disturbing F-actin distribution and localization of multiple actin-binding proteins. Cre-mediated deletion of Cttn also did not affect cell migration. To obtain mice with a Cttn null allele, we next crossed Cttnflox/flox mice with transgenic mice that express Cre recombinase ubiquitously. Western blot and immunocytochemical analysis confirmed complete elimination of cortactin expression in MEFs carrying homozygously Cttn null alleles. Howev... genesis journals http://www.medworm.com/rss/comments.php?id=2619624 Sun, 19 Jul 2009 23:00:00 +0100 2619624 http://www.medworm.com/index.php?rid=2619623&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20547 The Hox family of transcription factors are expressed at different domains along the rostrocaudal (R-C) body axis during development. To examine the function of Hoxc8 and Hoxc9 in specific cell types and at different developmental times, we have generated and characterized loxP flanked (floxed) Hoxc8 and Hoxc8[rarr]Hoxc9 replacement alleles of mice, with either GFP or LacZ reporters. Although all four alleles of mice behave like wild-type controls in motor behavioral testing, slight differences in endogenous Hox gene expression were observed among these alleles depending on the type of reporters used and the presence of Hoxc9 cDNA in the targeting constructs. The efficiency of Cre-mediated recombination was evaluated by crossing these mice with the Nestin-cre and Isl1-cre mice, and the los... genesis journals http://www.medworm.com/rss/comments.php?id=2619623 Sun, 19 Jul 2009 23:00:00 +0100 2619623 http://www.medworm.com/index.php?rid=2619622&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20548 Phosphorylation of Ser133 in the transcription factor CREB is an important mechanism for regulating its transcriptional activity, however recent work has suggested significant roles for other regulatory inputs into CREB. To allow study of this in vivo, we have generated a Ser133 to alanine knockin mutation in the mouse CREB locus. As CREB knockout is perinatal lethal, a minigene strategy was used to allow conditional knockin of the Ser133Ala mutation in adult mice using Cre recombinase. While some expression of the mutated protein was observed prior to Cre expression, following Cre expression in either T cells or neurons only the mutated CREB protein was detected. genesis 00:1-9, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2619622 Sun, 19 Jul 2009 23:00:00 +0100 2619622 http://www.medworm.com/index.php?rid=2619621&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20549 In conclusion, the LC-1-hCD19-CreERT2 strain is a powerful tool to study gene function specifically in B lymphocytes at any chosen time point in the lifecycle of the mouse. genesis 00:1-7, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2619621 Sun, 19 Jul 2009 23:00:00 +0100 2619621 http://www.medworm.com/index.php?rid=2619620&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20551 Consistent left-right patterning is a fascinating and biomedically important problem. In the chick embryo, it is not known how cells determine their position (left or right) relative to the primitive streak, which is required for subsequent asymmetric gene expression cascades. We show that the subcellular localization of Vangl2, a core planar cell polarity (PCP) protein, is consistently polarized, giving cells in the blastoderm a vector pointing toward the primitive streak. Moreover, morpholino-mediated loss-of-function of Vangl2 by electroporation into chicks at very early stages randomizes the normally left-sided expression of Sonic hedgehog. Strikingly, Vangl2 morpholinos also induce a desynchronization of asymmetric gene expression within the left and right domains of Hensen's node. Th... genesis journals http://www.medworm.com/rss/comments.php?id=2619620 Sun, 19 Jul 2009 23:00:00 +0100 2619620 http://www.medworm.com/index.php?rid=2600491&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20552 Gastric pit cells are high-turnover epithelial cells of the gastric mucosa. They secrete mucus to protect the gastric epithelium from acid and pepsin. To investigate the genetic mechanisms underlying the physiological functions of gastric pit cells, we generated a transgenic mouse line, namely, Capn8-Cre, in which the expression of Cre recombinase was controlled by the promoter of the intracellular Ca2+-regulated cysteine protease calpain-8. To test the tissue distribution and excision activity of Cre recombinase, the Capn8-Cre transgenic mice were bred with the ROSA26 reporter strain and a mouse strain that carries Smad4 conditional alleles (Smad4Co/Co). Multiple-tissue PCR and LacZ staining demonstrated that Capn8-Cre transgenic mouse expressed Cre recombinase in the gastric pit cells. C... genesis journals http://www.medworm.com/rss/comments.php?id=2600491 Tue, 14 Jul 2009 23:00:00 +0100 2600491 http://www.medworm.com/index.php?rid=2600496&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20534 Each cell within a multicellular organism has distinguishable characteristics established by its unique patterns of gene expression. This individual identity is determined by the expression of genes in a time and place-dependent manner, and it is becoming increasingly clear that chromatin plays a fundamental role in the control of gene transcription in multicellular organisms. Therefore, understanding the regulation of chromatin and how the distinct identity of a cell is passed to daughter cells during development is paramount. Techniques with which to study chromatin have advanced rapidly over the past decade. Development of high throughput techniques and their proper applications has provided us essential tools to understand the regulation of epigenetic phenomena and its effect on gene e... genesis journals http://www.medworm.com/rss/comments.php?id=2600496 Mon, 13 Jul 2009 23:00:00 +0100 2600496 http://www.medworm.com/index.php?rid=2600495&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20535 GABAB receptors are the G-protein-coupled receptors for the neurotransmitter [gamma]-aminobutyric acid (GABA). Receptor subtypes are based on the subunit isoforms GABAB1a and GABAB1b, which combine with GABAB2 subunits to form heteromeric receptors. Here, we used a modified bacterial artificial chromosome (BAC) containing the GABAB1 gene to generate transgenic mice expressing GABAB1a and GABAB1b subunits fused to the enhanced green fluorescence protein (eGFP). We demonstrate that the GABAB1-eGFP fusion proteins reproduce the cellular expression patterns of endogenous GABAB1 proteins in the brain and in peripheral tissue. Crossing the GABAB1-eGFP BAC transgene into the GABAB1-/- background restores pre and postsynaptic GABAB functions, showing that the GABAB1-eGFP fusion proteins substitute... genesis journals http://www.medworm.com/rss/comments.php?id=2600495 Mon, 13 Jul 2009 23:00:00 +0100 2600495 http://www.medworm.com/index.php?rid=2600494&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20536 The classical cell sorting experiments undertaken by Townes and Holtfreter described the intrinsic propensity of dissociated embryonic cells to self-organize and reconcile into their original embryonic germ layers with characteristic histotypic positioning. Steinberg presented the differential adhesion hypothesis to explain these patterning phenomena. Here, we have reappraised these issues by implementing embryoid bodies to model the patterning of epiblast and primitive endoderm layers. We have used combinations of embryonic stem (ES) cells and their derivatives differentiated by retinoic acid treatment to model epiblast and endoderm cells, and wild-type or E-cadherin null cells to represent strongly or weakly adherent cells, respectively. One cell type was fluorescently labeled and recons... genesis journals http://www.medworm.com/rss/comments.php?id=2600494 Mon, 13 Jul 2009 23:00:00 +0100 2600494 http://www.medworm.com/index.php?rid=2600493&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20545 We report the generation of a bioluminescent reporter strain that allows monitoring of tumor development in preexisting conditional mouse tumor models. We demonstrate that, in a Cre-dependent glioblastoma multiforme model, tumor initiation and progression is readily monitored over time and that luminescent output is related to tumor volume. Our results show that this reporter strain may be combined with various Cre/loxP mouse tumor models to allow for noninvasive longitudinal monitoring of tumor growth and therapeutic response in vivo. genesis 00:1-8, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2600493 Mon, 13 Jul 2009 23:00:00 +0100 2600493 http://www.medworm.com/index.php?rid=2600492&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20546 We describe here a conditional Gbx2 gain-of-function transgenic mouse line, Gbx2-GOF, which expresses Gbx2 and red fluorescence protein, mCherry, upon Cre-mediated recombination. In the absence of Cre, [beta]-galactosidase is broadly expressed in mouse embryos and adult brains carrying the transgene. By combining Gbx2-GOF and En1Cre knock-in allele, we activated expression of Gbx2 and mCherry throughout the mesencephalon (mes) and rhombomere 1 (r1). The ectopic expression of Gbx2 causes an anterior shift of the mes/r1 junction at embryonic day 10.5. Interestingly, we found that persistent expression of Gbx2 throughout the mes/r1 region largely abolishes expression of the isthmic organizer gene Fgf8, leading to deletion of the midbrain and cerebellum at later stages. Our data suggest that t... genesis journals http://www.medworm.com/rss/comments.php?id=2600492 Mon, 13 Jul 2009 23:00:00 +0100 2600492 http://www.medworm.com/index.php?rid=2500592&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20540 Sox17 encodes an SRY-related high-mobility group (HMG) box transcription factor that is essential for endoderm formation and fetal hematopoietic stem cell maintenance. In the mouse, expression of Sox17 is first observed in the extraembryonic endoderm and is subsequently seen in the definitive endoderm as well as in blood and the endothelial cells of the developing vasculature. To conditionally inactivate genes in these domains, we have targeted the Sox17 locus to generate a bicistronic mRNA linking Sox17 to a codon improved Cre recombinase (iCre) via a viral 2A sequence. Here we report a new Cre knock-in mouse line, Sox17-2A-iCre, with activity in the developing endoderm, the vascular endothelial cells of the cardiovascular system and the hematopoietic system. Our results indicate that the... genesis journals http://www.medworm.com/rss/comments.php?id=2500592 Wed, 24 Jun 2009 23:00:00 +0100 2500592 http://www.medworm.com/index.php?rid=2500594&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20538 Glucose-6-phosphatase-[alpha] (G6Pase-[alpha] or G6PC) catalyzes the hydrolysis of glucose-6-phosphate to glucose and is a key enzyme in interprandial glucose homeostasis. Mutations in the human G6PC gene, expressed primarily in the liver, kidney, and intestine, cause glycogen storage disease Type Ia (GSD-Ia), an autosomal recessive disorder characterized by a disturbed glucose homeostasis. For better understanding of the roles of G6Pase-[alpha] in different tissues and in pathological conditions, we have generated mice harboring a conditional null allele for G6pc by flanking Exon 3 of the G6pc gene with loxP sites. We confirmed the null phenotype by using the EIIa-Cre transgenic approach to generate mice lacking Exon 3 of the G6pc gene. The resulting homozygous Cre-recombined null mice ma... genesis journals http://www.medworm.com/rss/comments.php?id=2500594 Sun, 21 Jun 2009 23:00:00 +0100 2500594 http://www.medworm.com/index.php?rid=2500593&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20539 Germ cells are essential for the propagation of individual species. Studies on germ cell development in mice highlight important biological paradigms. Beginning with their first appearance around embryonic day 7 (E7), germ cells undergo specific cellular changes at different stages of their embryonic and adult development. Germ cells migrate through the hind-regions of the embryo to eventually home into the developing gonads. Further differentiation and development of germ cells differ in males and females. The processes involved in germ cell development and their eventual differentiation into sperm and oocytes have been under extensive investigation in recent years. Studies on germ cells have shed light on the cellular and molecular processes involved in their specification, migration, pr... genesis journals http://www.medworm.com/rss/comments.php?id=2500593 Sun, 21 Jun 2009 23:00:00 +0100 2500593 http://www.medworm.com/index.php?rid=2500596&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20532 Development of the mammalian adrenal gland is regulated by a diverse network of growth and transcription factors. Disruptions in these pathways often result in adrenal insufficiency because of adrenal hypoplasia. Several lines of evidence have suggested that the Hedgehog signaling pathway, which controls many aspects of tissue and organ patterning, may play a direct role in adrenal morphogenesis as well. Therefore, we examined the role of Sonic Hedgehog (Shh), a member of the Hedgehog family, in mouse adrenal development. We show that Shh and its downstream effectors Gli1, Gli2, and Gli3 are expressed in the adrenal cortex throughout development, and that Shh is required for normal adrenal organogenesis. Conditional inactivation of Shh in the adrenal cortex using a Cre-loxP system resulted... genesis journals http://www.medworm.com/rss/comments.php?id=2500596 Tue, 16 Jun 2009 23:00:00 +0100 2500596 http://www.medworm.com/index.php?rid=2500595&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20543 Repression by E(spl)M8 during inhibitory Notch (N) signaling (lateral inhibition) is regulated, in part, by protein kinase CK2, but the involvement of a phosphatase has been unclear. The studies we report here employ Tik, a unique dominant-negative (DN) mutation in the catalytic subunit of CK2, in a Gal4-UAS based assay for impaired lateral inhibition. Specifically, overexpression of Tik elicits ectopic bristles in N+ flies and suppresses the retinal defects of the gain-of-function allele Nspl. Functional dissection of the two substitutions in Tik (M161K and E165D), suggests that both mutations contribute to its DN effects. While the former replacement compromises CK2 activity by impairing ATP-binding, the latter affects a conserved motif implicated in binding the phosphatase PP2A. Accordi... genesis journals http://www.medworm.com/rss/comments.php?id=2500595 Tue, 16 Jun 2009 23:00:00 +0100 2500595 http://www.medworm.com/index.php?rid=2500598&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20537 In this report we describe the construction of a transgenic mouse strain that expresses the Notch2 intracellular domain in response to cell lineage specific expression of Cre recombinase. This approach bypasses the requirement for ligand- receptor interaction and allows the direct determination of the consequences of Notch2 activation in vivo. Exogenous expression of the Notch2 intracellular domain resulted in the developmental arrest of secondary heart field derived cardiomyocytes during the transition from immature [alpha]-Smooth Muscle Actin expressing cells to mature [alpha]-Actinin positive cardiomyocytes. In contrast, a cell nonautonomous mesenchymal expansion was observed in semilunar valves. This new conditionally expressed allele of Notch2 can be used in studies by investigators i... genesis journals http://www.medworm.com/rss/comments.php?id=2500598 Sun, 14 Jun 2009 23:00:00 +0100 2500598 http://www.medworm.com/index.php?rid=2500597&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20541 In this study, we first revealed the allelic enrichment of histone modifications and variant histones at the imprinting control regions (ICRs) of four-cell to blastocyst stage preimplantation embryos by using carrier chromatin immunoprecipitation and sequence polymorphism analysis of immunoprecipitated DNA. We found relative enrichment of histone H3 lysine 9 dimethylation at the imprinted alleles of ICRs and obtained the results suggesting that histone modifications at ICRs are established during a late preimplantation stage. genesis, 00:1-6, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2500597 Sun, 14 Jun 2009 23:00:00 +0100 2500597 http://www.medworm.com/index.php?rid=2500599&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20531 The chemokine receptors CCR2 and CCR5 represent potential novel therapeutic targets to treat important inflammatory and infectious diseases, including atherosclerosis and HIV infection. To study the functions of both receptors in vivo, we aimed to generate Ccr2/Ccr5 double-deficient mice. As these genes are separated by (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2500599 Tue, 09 Jun 2009 23:00:00 +0100 2500599 http://www.medworm.com/index.php?rid=2411691&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20518 In this report, we have adapted a lentiviral gene delivery technique for genetic modification of the rat trophoblast cell lineage. Blastocysts were incubated with lentiviral particles and transferred into the uteri of pseudopregnant female rats, harvested at various times during gestation, and then analyzed. Two test systems were evaluated: (1) delivery of an enhanced green fluorescent protein (EGFP) gene under the control of constitutive promoters to rat blastocysts; (2) delivery of EGFP short hairpin RNA (shRNA) to rat blastocysts constitutively expressing EGFP. Lentiviral packaged gene constructs were efficiently and specifically delivered to all trophoblast cell lineages. Additionally, lentiviral mediated transfer of shRNAs was an effective strategy for modifying gene expression in tro... genesis journals http://www.medworm.com/rss/comments.php?id=2411691 Fri, 15 May 2009 04:00:00 +0100 2411691 http://www.medworm.com/index.php?rid=2395686&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20528 MAM domain containing glycosilphosphatidilinositol anchor 1 (MDGA1) is an IgCAM protein present in many vertebrate species including humans. In mammals, MDGA1 is expressed by a subset of neurons in the developing brain and thought to function in neural cell migration. We identified a fish ortholog of mdga1 by a gene-trap screen utilizing the Frog Prince transposon in medaka (Japanese killifish, Oryzias latipes). The gene-trap vector was inserted into an intronic region of mdga1 to form a chimeric protein with green fluorescent protein, allowing us to monitor mdga1 expression in vivo. Expression of medaka mdga1 was seen in various types of embryonic brain neurons, and specifically in neurons migrating toward their target sites, supporting the proposed function of MDGA1. We also isolated the... genesis journals http://www.medworm.com/rss/comments.php?id=2395686 Thu, 07 May 2009 04:00:00 +0100 2395686 http://www.medworm.com/index.php?rid=2395689&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20517 The Sine Oculis Homeobox (SIX) proteins play critical roles in organogenesis and are defined by the presence of two evolutionarily conserved functional motifs: a homeobox DNA binding domain and the SIX protein-protein interaction domain. Members of this transcription factor family can be divided into three subgroups: Six1/2, Six4/5, and Six3/6. This partitioning is based mainly on protein sequence similarity and genomic architecture, and not on specificities of DNA binding or binding partners. In fact, it is well demonstrated that members of the different subgroups can bind to and activate common transcriptional targets as well as form biochemical complexes with communal binding partners. Here we report that the C-terminal segment, which is not conserved across different SIX subfamilies, m... genesis journals http://www.medworm.com/rss/comments.php?id=2395689 Wed, 06 May 2009 04:00:00 +0100 2395689 http://www.medworm.com/index.php?rid=2395688&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20529 Mahogunin Ring Finger-1 (Mgrn1) null mutant mice have a pleiotropic phenotype that includes the absence of yellow hair pigment, abnormal head shape, reduced viability, and adult-onset spongiform neurodegeneration. Mgrn1 encodes a highly conserved E3 ubiquitin ligase with four different isoforms which are differentially expressed and predicted to localize to different subcellular compartments. To test whether loss of specific isoforms causes different aspects of the mutant phenotype, we generated transgenes for each isoform and bred them onto the null mutant background. Mice expressing only isoform I or III appeared completely normal. Isoform II rescued or partially rescued the mutant phenotypes, whereas isoform IV had little or no effect. Our data show that different Mgrn1 isoforms are not... genesis journals http://www.medworm.com/rss/comments.php?id=2395688 Wed, 06 May 2009 04:00:00 +0100 2395688 http://www.medworm.com/index.php?rid=2395687&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20530 Phytophthora capsici causes serious diseases in numerous crop plants. Polygalacturonases (PGs) are cell wall-degrading enzymes that play an important role in pathogenesis in straminopilous pathogens. To understand PGs as they relate to the virulence of P. capsici, Pcipg2 was identified from a genomic library of a highly virulent P. capsici strain. Pcipg2 was strongly expressed during symptom development after the inoculation of pepper leaves with P. capsici. The wild protein (PCIPGII) was obtained from the expression of pcipg2 and found that increasing activity of PGs in PCIPGII-treated pepper leaves was consistent with increasing symptom development. Asp residues in active sites within pcipg2 affected PCIPGII activity or its virulence on pepper leaves. Results show that pcipg2 is an impor... genesis journals http://www.medworm.com/rss/comments.php?id=2395687 Wed, 06 May 2009 04:00:00 +0100 2395687 http://www.medworm.com/index.php?rid=2395697&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20515 Tbx18 is a T-Box transcription factor that has specific expression and indispensible function in the lower urinary tract. Here, we report the generation and characterization of a bacterial artificial chromosome (BAC) transgene expressing Cre under the control of Tbx18 regulatory elements. When crossed to the ROSA26R-lacZ reporter mice, the Tbx18-Cre transgene mediates loxP recombination in the mesenchymal derivatives in the lower urinary tract, especially in the smooth muscle cells (SMCs) and the stromal cells. There is no expression of this transgene in the urothelium or in the kidney. This Tbx18-Cre transgene recapitulates the endogenous Tbx18 expression in the urinary system and can be used for the study of the development, physiology, and diseases in the urinary tract. Its additional e... genesis journals http://www.medworm.com/rss/comments.php?id=2395697 Mon, 04 May 2009 04:00:00 +0100 2395697 http://www.medworm.com/index.php?rid=2395696&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20516 Transforming growth factor [beta]1 (TGF[beta]1) is a multifunctional growth factor involved in wound healing, tissue fibrosis, and in the pathogenesis of many syndromic diseases (e.g., Marfan syndrome, Camurati-Engelmann disease) and muscular, neurological, ophthalmic, cardiovascular and immunological disorders, and cancer. Since the generation of Tgfb1 knockout mice, there has been extraordinary progress in understanding its physiological and pathophysiological function. Here, we report the generation of a conditional knockout allele for Tgfb1 in which its exon 6 is flanked with LoxP sites. As proof of principle, we crossed these mice to LckCre transgenic mice and specifically disrupted Tgfb1 in T cells. The results indicate that T-cell-produced TGF[beta]1 is required for normal in vivo r... genesis journals http://www.medworm.com/rss/comments.php?id=2395696 Mon, 04 May 2009 04:00:00 +0100 2395696 http://www.medworm.com/index.php?rid=2395695&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20520 The HMG-box transcription factor Sox17 has been shown to play important roles in both endoderm formation and cardiovascular development. To conditionally inactivate genes in these domains, we have targeted a codon improved Cre Recombinase (iCre) into exon 1 of the Sox17 gene. Surprisingly, Cre-mediated recombination in the Rosa26 reporter mouse line revealed largely specific activity within the vasculature rather than in endoderm-derived tissues. Here we report a new Cre knock-in mouse line, Sox17iCre with activity in the vascular endothelial cells of arteries in the cardiovascular system but not in veins. Cre-mediated recombination was also strongly detected in the liver and spleen, the two organs associated with hematopoiesis. Thus, these results indicate that the Sox17iCre would be an a... genesis journals http://www.medworm.com/rss/comments.php?id=2395695 Mon, 04 May 2009 04:00:00 +0100 2395695 http://www.medworm.com/index.php?rid=2395694&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20522 Transthyretin (Ttr) is a thyroid hormone transport protein secreted by cells of the visceral yolk sac and fetal liver in developing embryos, and by hepatocytes and the choroid plexus epithelium of the brain in adult mice. Spatiotemporal localization of Ttr mRNA during embryogenesis suggested that Ttr regulatory elements might drive transgene expression throughout the visceral endoderm of early mouse embryos. We use Ttr cis-regulatory elements to generate Ttr::RFP and Ttr::Cre strains of mice, driving red fluorescent protein (RFP) and a nuclear-localized Cre recombinase, respectively. Visualization of RFP fluorescence in Ttr::RFP transgenics confirms reporter localization throughout the visceral endoderm in early embryos and in the visceral yolk sac and fetal liver of later stage embryos. U... genesis journals http://www.medworm.com/rss/comments.php?id=2395694 Mon, 04 May 2009 04:00:00 +0100 2395694 http://www.medworm.com/index.php?rid=2395693&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20524 Smad7 can be induced by various transforming growth factor-[beta] superfamily ligands and negatively modulates their signaling, thus acting in a negative, autocrine feedback manner. Previous analyses have demonstrated that although Smad7 is widely expressed, it is predominantly found in the vascular endothelium. Because of the restricted spatiotemporal reporter expression driven via a novel 4.3 kb Smad7 promoter in endocardial cells overlying the hearts atrioventricular (AV) cushions; we hypothesized that a transgenic Cre line would prove useful for the analysis of endocardial cushion and valve formation. Here we describe a mouse line, Smad7Cre, where Cre is robustly expressed within both cardiac outflow and AV endocardial cushions. Additionally, as endocardial cells are thought to contrib... genesis journals http://www.medworm.com/rss/comments.php?id=2395693 Mon, 04 May 2009 04:00:00 +0100 2395693 http://www.medworm.com/index.php?rid=2395692&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20521 Our results, using endogenous mutants and Gal4-UAS driven transgenes, implicate multisite phosphorylation in repression by E(spl)M8. We propose that these phosphorylations occur in the morphogenetic furrow (MF) to reverse an auto-inhibited state of M8, enabling repression of Atonal during R8 specification. Our studies address the paradoxical behavior of M8*, the truncated protein encoded by E(spl)D. We suggest that differences in N signaling in the bristle versus the eye underlie the antimorphic activity of M8* in N+ (ectopic bristles) and hypermorphic activity in Nspl (reduced eye). Ectopic M8* impairs eye development (in Nspl) only during establishment of the atonal feedback loop (anterior to the MF), but is ineffective after this time point. In contrast, a CK2 phosphomimetic M8 lacking ... genesis journals http://www.medworm.com/rss/comments.php?id=2395692 Mon, 04 May 2009 04:00:00 +0100 2395692 http://www.medworm.com/index.php?rid=2395691&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20526 Site-specific recombinases such as Cre and Flp are invaluable tools for genetic manipulations, but their usage in zebrafish has been limited. Incorporating recently developed flip-excision (FlEx) design that allows stable inversions, we have established zebrafish reporter lines that express bright and ubiquitous EGFP, but switch to express mCherry in the presence of Cre or Flp. Here, we demonstrate the stable inversion in the reporter lines, both in somatic cells and in the germ line by Cre or Flp, and the subsequent reinversion using the other recombinase. Using the reporter lines, we characterized cardiomyocyte-specific Cre lines and neuronal progenitor-specific and tamoxifen-dependent Cre lines. We also used the reporter lines for screening Cre- and Flp-based enhancer trap lines. Simila... genesis journals http://www.medworm.com/rss/comments.php?id=2395691 Mon, 04 May 2009 04:00:00 +0100 2395691 http://www.medworm.com/index.php?rid=2395690&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20527 The Drosophila roughest (rst) locus encodes an immunoglobulin superfamily transmembrane glycoprotein implicated in a variety of embryonic and postembryonic developmental processes. Here we demonstrate a previously unnoticed role for this gene in the autophagic elimination of larval salivary glands during early pupal stages by showing that overexpression of the Rst protein ectodomain in early pupa leads to persistence of salivary glands up to at least 12 hours after head eversion, although with variable penetrance. The same phenotype is observed in individuals carrying the dominant regulatory allele rstD, but not in loss of function alleles. Analysis of persistent glands at the ultrastructural level showed that programmed cell death starts at the right time but is arrested at an early stage... genesis journals http://www.medworm.com/rss/comments.php?id=2395690 Mon, 04 May 2009 04:00:00 +0100 2395690 http://www.medworm.com/index.php?rid=2363007&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20519 ROCK kinases regulate actin-myosin structures downstream of Rho GTPases. We generated mice expressing 4-hydroxytamoxifen (4HT)-regulated human ROCK II (ROCKII:mERTM) under the transcriptional control of the cytokeratin14 (K14) promoter. The K14-ROCKII:mERTM minigene was recombineered into a novel cloning vector containing the promoter and first exon of the human HPRT gene, and second and third exons of the mouse Hprt gene. Homologous recombination into the Hprt locus, which is deleted for the promoter and first two exons in HM1 embryonic stem cells, reconstitutes a functional Hprt gene, allowing for growth in HAT (hypoxanthine-aminopterin-thymidine) medium. K14-promoter-driven ROCKII:mERTM expression was restricted to a superficial cell layer in embryoid bodies, with increased ROCK substra... genesis journals http://www.medworm.com/rss/comments.php?id=2363007 Fri, 24 Apr 2009 04:00:00 +0100 2363007 http://www.medworm.com/index.php?rid=2363012&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20508 The Sleeping Beauty and PiggyBac DNA transposon systems have recently been developed as tools for insertional mutagenesis. We have compared the chromosomal mobilization efficiency and insertion site preference of the two transposons mobilized from the same donor site in mouse embryonic stem (ES) cells under conditions in which there were no selective constraints on the transposons' insertion sites. Compared with Sleeping Beauty, PiggyBac exhibits higher transposition efficiencies, no evidence for local hopping and a significant bias toward reintegration in intragenic regions, which demonstrate its utility for insertional mutagenesis. Although Sleeping Beauty had no detectable genomic bias with respect to insertions in genes or intergenic regions, both Sleeping Beauty and PiggyBac transposo... genesis journals http://www.medworm.com/rss/comments.php?id=2363012 Thu, 23 Apr 2009 04:00:00 +0100 2363012 http://www.medworm.com/index.php?rid=2363011&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20509 In humans mutations in DKC1, cause the rare bone marrow failure syndrome dyskeratosis congenita. We have used gene targeting to produce mouse ES cells with Dkc1 mutations that cause DC when in humans. The mutation A353V, the most common human mutation, causes typical DC to very severe DC in humans. Male chimeric mice carrying this mutation do not pass the mutated allele to their offspring. The mutation G402E accounts for a single typical case of DC in a human family. The allele carrying this mutation was transmitted to the offspring with high efficiency. Expression of RNA and protein was reduced compared to wild type animals, but no abnormalities of growth and development or in blood values were found in mutant mice. Thus Dkc1 mutations have variable expression inmice, as in humans. genesi... genesis journals http://www.medworm.com/rss/comments.php?id=2363011 Thu, 23 Apr 2009 04:00:00 +0100 2363011 http://www.medworm.com/index.php?rid=2363010&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20510 The piebald deletion complex is a set of overlapping chromosomal deficiencies on distal mouse chromosome 14. We surveyed the functional genetic content of the piebald deletion region in an essential gene mutagenesis screen of 952 genomes to recover seven lethal mutants. The ENU-induced mutations were mapped to define genetic intervals using the piebald deletion panel. Lethal mutations included loci required for establishment of the left-right embryonic axis and a loss-of-function allele of Phr1 resulting in respiratory distress at birth. A functional map of the piebald region integrates experimental genetic data from the deletion panel, mutagenesis screen, and the targeted disruption of specific genes. A comparison of several genomic intervals targeted in regional mutagenesis screens sugge... genesis journals http://www.medworm.com/rss/comments.php?id=2363010 Thu, 23 Apr 2009 04:00:00 +0100 2363010 http://www.medworm.com/index.php?rid=2363009&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20511 Bone morphogenetic proteins (BMPs) have multiple roles during embryogenesis. Current data indicate that the dosage of BMPs is tightly regulated for normal development in mice. Since Bmp2 or Bmp4 homozygous mutant mice show early embryonic lethality, we generated compound heterozygous mice for Bmp2 and Bmp4 to explore the impact of lowered dosage of these BMP ligands. Genotyping pups bred between Bmp2 and Bmp4 heterozygous mice revealed that the ratio of adult compound heterozygous mice for Bmp2 and Bmp4 is much lower than expected. During embryogenesis, the compound heterozygous embryos showed several abnormalities, including defects in eye formation, body wall closure defects, and ventricular septal defects (VSD) in the heart. However, the ratio of the compound heterozygous embryos was th... genesis journals http://www.medworm.com/rss/comments.php?id=2363009 Thu, 23 Apr 2009 04:00:00 +0100 2363009 http://www.medworm.com/index.php?rid=2363008&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20514 C57BL/6 (B6)-derived embryonic stem (ES) cells are not widely used to generate knockout mice despite the advantage of a well-defined genetic background because of poor developmental potential. We newly established serum- and feeder-free B6 ES cells with full developmental potential by using leukemia inhibitory factor (LIF) and 6-bromoindirubin-3[prime]-oxime (BIO), a glycogen synthase kinase-3 (GSK3) inhibitor. BIO treatment significantly increased the expression levels of 364 genes including pluripotency markers such as Nanog and Klf family. Unexpectedly, by aggregating or microinjecting those ES cells to each eight-cell-stage diploid embryo, we stably generated germline-competent ES-derived mice. Furthermore, founder mice completely derived from female XO, heterozygous, or homozygous mut... genesis journals http://www.medworm.com/rss/comments.php?id=2363008 Thu, 23 Apr 2009 04:00:00 +0100 2363008 http://www.medworm.com/index.php?rid=2338378&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20507 The rhombomere 4(r4)-restricted expression of the mouse Hoxb2 gene is regulated by a 1.4-kb enhancer-containing fragment. Here, we showthat transgenic mouse lines expressing cre driven by this fragment (B2-r4-Cre), activated the R26R Cre reporter in rhombomere 4 and the second branchial arch, the epithelium of the first branchial arch, apical ectodermal ridge of the limb buds and the tail region. Of particular interest is Cre activity in the developing inner ear. Cre activity was found in the preotic field and otic placode at E8.5 and otocyst at E9.5-E12.5, in the cochleovestibular and facio-acoustic ganglia at E10.5 and the vestibular and spiral ganglia and all the otic epithelia derived from the otocyst at E15.5 and P0. Our data suggest that the B2-r4-Cre transgenic mice provide an impor... genesis journals http://www.medworm.com/rss/comments.php?id=2338378 Sat, 18 Apr 2009 04:00:00 +0100 2338378 http://www.medworm.com/index.php?rid=2338380&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20505 Cellular reaggregation methods have been used to generate in vitro organotypic cultures as a means to elucidate the cellular and molecular requirements of organogenesis. However, reproducibility from experiment to experiment has remained problematic and furthermore, current protocols do not support reaggregation of many important tissues. Here, using the thymus as a model organ, we present a novel reaggregation method termed "compaction reaggregation" that offers improved kinetics of reaggregation and greatly improved efficiency. Using compaction reaggregation we have been able to reaggregate the aorta-gonad- mesonephros region, a tissue that previously proved refractory to commonly used reaggregation methods, enabling the study of hematopoietic stem cell emergence and expansion. Additiona... genesis journals http://www.medworm.com/rss/comments.php?id=2338380 Wed, 15 Apr 2009 04:00:00 +0100 2338380 http://www.medworm.com/index.php?rid=2338379&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20506 Homeobox gene Msx2 is widely expressed during both embryogenesis and postnatal development and plays important roles during organogenesis. We developed an Msx2-rtTA BAC transgenic line which can activate TetO-Cre expression in Msx2-expressing cells upon doxycycline (Dox) treatment. Using the Rosa26-LacZ (R26R) reporter line, we show that rtTA is activated in Msx2-expressing organs including the limb, heart, external genitalia, urogenital system, hair follicles and craniofacial regions. Moreover, we show that in body appendages, the transgene can be activated in different domains depending on the timing of Dox treatment. In addition, the transgene can also be effectively activated in adult tissues such as the hair follicle and the urogenital system. Taken together, this Msx2-rtTA;TetO-Cre s... genesis journals http://www.medworm.com/rss/comments.php?id=2338379 Wed, 15 Apr 2009 04:00:00 +0100 2338379 http://www.medworm.com/index.php?rid=2321118&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20504 Dogs (Canis familiaris) share many common genetic diseases with humans and development of disease models using a transgenic approach has long been awaited. However, due to the technical difficulty in obtaining fertilizable eggs and the unavailability of embryonic stem cells, no transgenic dog has been generated. Canine fetal fibroblasts were stably transfected with a red fluorescent protein (RFP) gene-expressing construct using retrovirus gene delivery method. Somatic cell nuclear transfer was then employed to replace the nucleus of an oocyte with the nucleus of the RFP-fibroblasts. Using this approach, we produced the first generation of transgenic dogs with four female and two male expressing RFP. genesis 00:1-9, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2321118 Fri, 10 Apr 2009 04:00:00 +0100 2321118 http://www.medworm.com/index.php?rid=2321123&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20500 This study, therefore, presents the feasibility of applying multiple spectrally and subcellularly distinct fluorescent protein reporters for live imaging studies in ES cells and mouse embryos. Furthermore, the increasing availability of spectral variant fluorescent proteins along with the development of methods that permit improved spectral separation now facilitate multiplexing of fluorescent reporters to provide readouts of a variety of anatomical and physiological behaviors simultaneously in living specimens. genesis 00:1-7, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2321123 Wed, 08 Apr 2009 04:00:00 +0100 2321123 http://www.medworm.com/index.php?rid=2321122&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20502 The synaptic vesicle protein, synaptotagmin I, is a multifunctional protein required for several steps in the synaptic vesicle cycle. It is primarily composed of two calcium-binding domains, C2A and C2B. Within each of these domains, a polylysine motif has been identified that is proposed to mediate specific functions within the synaptic vesicle cycle. While the C2B polylysine motif plays an important role in synaptic transmission in vivo, the C2A polylysine motif has not previously been analyzed at an intact synapse. Here, we show that mutation of the C2A polylysine motif increases the frequency of spontaneous transmitter release in vivo. The increased frequency is not a developmental consequence of disrupted synaptic transmission, as evoked transmitter release is unimpaired in the mutant... genesis journals http://www.medworm.com/rss/comments.php?id=2321122 Wed, 08 Apr 2009 04:00:00 +0100 2321122 http://www.medworm.com/index.php?rid=2321120&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20513 BMPRIA and its high-affinity ligand BMP4 have recently been shown to be expressed in the [beta]-cells of the pancreas. Here, we report the abnormalities of heterozygous mice for Bmpr1a in glucose metabolism during the course of intraperitoneal glucose tolerance test. The heterozygous mice had increased blood glucose levels throughout the first 2.5 h after the administration of glucose. Analysis of glucose-stimulated insulin secretion (GSIS) indicates that insulin secretion in the heterozygous mice is compromised, and induction of secreted insulin by stimulation is substantially lower compared with the wild-type controls. No apparent abnormalities in pancreas, thyroid, and liver were seen upon histological examination. Real-time PCR results of selected genes showed an increase in the mRNA l... genesis journals http://www.medworm.com/rss/comments.php?id=2321120 Wed, 08 Apr 2009 04:00:00 +0100 2321120 http://www.medworm.com/index.php?rid=2281677&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20503 To study paraxial mesoderm formation in the mouse, transgenic lines that can be used to either selectively delete or express genes of interest in the paraxial mesoderm are required. We have generated a transgenic mouse line that expresses Cre recombinase in the paraxial mesoderm (PAM) beginning at e7.5. A lacZ Cre recombinase reporter line showed that in addition to PAM and its derivatives, lateral plate and intermediate mesoderm derivatives were also exposed to Cre activity, while the node, notochord, and cardiac mesoderm were not. We further demonstrate that 70-75% of the fibroblasts generated from Dll1-msd Cre, ROSA26-rtTA embryos possess Cre recombinase activity. These mice can therefore be used in combination with tet-responsive transgenic lines to generate mesoderm-derived embryonic ... genesis journals http://www.medworm.com/rss/comments.php?id=2281677 Sat, 21 Mar 2009 04:00:00 +0100 2281677 http://www.medworm.com/index.php?rid=2281684&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20430 The LIM-homeodomain transcription factor lmx1b is a critical regulator of vertebrate dorsal-ventral limb patterning. In the mouse embryo, lmx1b is initially transcribed throughout most, if not all, limb bud cells at early stages, but then rapidly becomes restricted specifically to dorsal mesenchymal cells with a sharp boundary between the dorsal-positive and ventral-negative expression domains. How this expression pattern is initially established is not well understood, nor are mechanism(s) that maintain a sharp dorsal-ventral boundary between lmx1b expressing and nonexpressing cells. Here, we employ a genetic fate mapping approach to establish that the transition from a broad expression domain of lmx1b to a restricted dorsal domain of expression involves selective loss of lmx1b expression... genesis journals http://www.medworm.com/rss/comments.php?id=2281684 Thu, 19 Mar 2009 04:00:00 +0100 2281684 http://www.medworm.com/index.php?rid=2281683&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20498 In this study, we focused the gonadal sex differentiation process in O. luzonensis under different testis determining gene. First, we observed the gonadal development of O. luzonensis histologically. We then analyzed the expression of Sox9a2/Sox9b, Dmrt1, and Foxl2 during early development. Our results suggest that the sexual differentiation of germ cells in O. luzonensis is initiated later than in O. latipes. However, the timing of the sexual differentiation of the supporting cell linage is similar between the species. genesis 00:1-11, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2281683 Thu, 19 Mar 2009 04:00:00 +0100 2281683 http://www.medworm.com/index.php?rid=2281681&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20499 We report the molecular cloning of a dominant Roller mutant of Pristionchus pacificus, which encodes a cuticle collagen. We use the mutant locus as a marker to develop transgenic technique by generating complex arrays and present flourescent-protein based transcriptional reporter constructs for P. pacificus. genesis 00:1-5, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2281681 Thu, 19 Mar 2009 04:00:00 +0100 2281681 http://www.medworm.com/index.php?rid=2281679&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20501 Ends-in and ends-out gene replacement approaches have been successfully used to disrupt Drosophila genes involved in a variety of biological processes. These methods combine double-strand breaks and homologous recombination to replace a targeted chromosome region with a designed DNA sequence. Unfortunately, these methods require large numbers of single animal crosses, making them both time consuming and labor intensive. Here, we designed a single complete targeting vector for use in a mass crossing ends-out gene targeting study. Importantly, our gene targeting method included a balancer chromosome to block endogenous homologous chromosome pairing and to promote pairing between the foreign targeting DNA fragment and the targeted chromosome. This technique provided successful and efficient g... genesis journals http://www.medworm.com/rss/comments.php?id=2281679 Thu, 19 Mar 2009 04:00:00 +0100 2281679 http://www.medworm.com/index.php?rid=2237120&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20468 The nuclear factor [kappa]B (NF-[kappa]B) signaling pathway regulates immune and inflammatory responses and is implicated in the pathogenesis of multiple diseases. The principal mechanism controlling NF-[kappa]B activation depends on the association of NF-[kappa]B transcription factor dimers with I[kappa]B molecules, which prevents the accumulation of NF-[kappa]B in the nucleus and the activation of target gene transcription. Monitoring the nucleocytoplalsmic shuttling of NF-[kappa]B factors is a reliable method to study the dynamic regulation of NF-[kappa]B activity. Here, we generated knock-in mice expressing a fusion protein between the green fluorescent protein (GFP) and the p65/RelA NF-[kappa]B subunit (GFP-p65) from the endogenous p65 genomic locus. Homozygous GFP-p65 mice developed ... genesis journals http://www.medworm.com/rss/comments.php?id=2237120 Fri, 06 Mar 2009 05:00:00 +0100 2237120 http://www.medworm.com/index.php?rid=2237121&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20497 IL-7 is a cytokine that is required for T-cell development and homeostasis as well as for lymph node organogenesis. Despite the importance of IL-7 in the immune system and its potential therapeutic relevance, questions remain regarding the sites of IL-7 synthesis, specific cell types involved and molecular mechanisms regulating IL-7 expression. To address these issues, we generated two bacterial artificial chromosome (BAC) transgenic mouse lines in which IL-7 regulatory elements drive expression of either Cre recombinase or a human CD25 (hCD25) cell surface reporter molecule. Expression of the IL-7.hCD25 BAC transgene, detected by reactivity with anti-hCD25 antibody, mimicked endogenous IL-7 expression. Fetal and adult tissues from crosses between IL-7.Cre transgenic mice and Rosa26R or R2... genesis journals http://www.medworm.com/rss/comments.php?id=2237121 Wed, 04 Mar 2009 05:00:00 +0100 2237121 http://www.medworm.com/index.php?rid=2219737&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20476 Mammalian development is highly sensitive to Tbx1 gene dosage reduction. Gene function insights can also be learned from increased or ectopic expression. The authors generated a novel mouse transgenic line, named COET, which expresses Tbx1 upon Cre-mediated recombination. The authors crossed this transgenic line with Tbx1Cre animals to activate expression in the Tbx1-expression domain. Compound mutant COET;Tbx1Cre/+ animals died after birth and showed heart enlargement. At E18.5, compound mutants showed ventricular septal defects and thymic abnormalities. The authors crossed compound mutants into a Tbx1 null background to understand whether this phenotype is caused by gene overdosage. Results showed that gene dosage reduction at the endogenous locus could not rescue heart and thymic defect... genesis journals http://www.medworm.com/rss/comments.php?id=2219737 Fri, 27 Feb 2009 05:00:00 +0100 2219737 http://www.medworm.com/index.php?rid=2213822&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20486 Ligands of the transforming growth factor-[beta] (TGF-[beta]) superfamily play important roles in embryonic patterning and development throughout the animal kingdom. Consequently, extracellular factors that affect ligand stability, mobility, and receptor interaction also have profound effects on development. One such regulator, Follistatin (Fst), functions as an inhibitor of both activin and bone morphogenetic protein (BMP) subfamilies of TGF-[beta] ligands in vertebrates. Drosophila follistatin (fs) encodes a Fst homolog that is broadly expressed throughout development, but the in vivo function of the protein remains unclear. We show that overexpression of fs affects prepupal to pupal transition and morphogenesis, highlighting a novel requirement for TGF-[beta] signaling in metamorphosis.... genesis journals http://www.medworm.com/rss/comments.php?id=2213822 Thu, 26 Feb 2009 05:00:00 +0100 2213822 http://www.medworm.com/index.php?rid=2213828&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20451 Mammalian androgenetic embryos can be produced by pronuclear exchange of fertilized oocytes or by dispermic in vitro fertilization of enucleated oocytes. Here, we report a new technique for producing mouse androgenetic embryos by injection of two round spermatid nuclei into oocytes, followed by female chromosome removal. We found that injection of round spermatids resulted in high rates of oocyte survival (88%). Androgenetic embryos thus produced developed into mid-gestation fetuses at various rates, depending on the mouse strain used. All the fetuses examined maintained paternally specific genomic imprinting memories. This technique also enabled us to produce complete heterozygous F1 embryos by injecting two spermatids from different strains. The best rate of fetal survival (12% per embry... genesis journals http://www.medworm.com/rss/comments.php?id=2213828 Mon, 23 Feb 2009 05:00:00 +0100 2213828 http://www.medworm.com/index.php?rid=2213827&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20472 Cajal-Retzius cells are an enigmatic class of neurons located in the most superficial layer of the cerebral cortex, and they play an important role in cortical development. Although many studies have indicated that CR cells are involved in regulating cell migration and cortical maturation, the function of these cells is still not fully understood. Here we describe an inducible Cre mouse line in which CreERTM is driven by the promoter for the Wnt receptor Frizzled10. Consistent with our previous studies on Frizzled10 expression and transgenic mouse lines using the Frizzled10 promoter, we found that in the developing telencephalon, Cre was mainly detected at the cortical hem, the largest source of CR cells. By crossing the Cre line to R26R reporter mice and injecting tamoxifen at different t... genesis journals http://www.medworm.com/rss/comments.php?id=2213827 Mon, 23 Feb 2009 05:00:00 +0100 2213827 http://www.medworm.com/index.php?rid=2213826&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20483 Malaria kills a million people annually. Understanding the relationship between a causative parasite, Plasmodium falciparum, and the mosquito vector might suggest novel prevention approaches. We created and transformed into Drosophila two genes encoding, thrombospondin-related adhesive protein (TRAP) and circumsporozoite protein (CSP), found on the cell surface of Plasmodium sporozoites. To understand a model insect's response, we induced these proteins separately and together, performing whole genome microarray analysis measuring gene expression changes. Gene ontology classification of responding genes reveals that TRAP and CSP strongly and differentially influence Drosophila genes involved with cell motility and gene regulation, respectively; however, the most striking effects are on the... genesis journals http://www.medworm.com/rss/comments.php?id=2213826 Mon, 23 Feb 2009 05:00:00 +0100 2213826 http://www.medworm.com/index.php?rid=2213825&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20484 Transgenic mice with a defined cell- or tissues-specific expression of Cre-recombinase are essential tools to study gene function. Here we report the generation and analysis of a transgenic mouse line (Cdx1::Cre) with restricted Cre-expression from Cdx1 regulatory elements. The expression of Cre-recombinase mimicked the endogenous expression pattern of Cdx1 at midgastrulation (from E7.5 to early headfold stage) inducing recombination in the three germlayers of the primitive streak region throughout the posterior embryo and caudal to the heart. This enables gene modifications to investigate patterning of the caudal embryo during and after gastrulation. Interestingly, we identified Cdx1 expression in the trophectoderm (TE) of blastocyst stage embryos. Concordantly, we detected extensive Cre-... genesis journals http://www.medworm.com/rss/comments.php?id=2213825 Mon, 23 Feb 2009 05:00:00 +0100 2213825 http://www.medworm.com/index.php?rid=2213824&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20485 The [gamma]-secretase complex is involved in cleaving transmembrane proteins such as Notch and one of the genes targeted in Alzheimer's disease known as amyloid precursor protein (APP). Presenilins function within the catalytic core of [gamma]-secretase, and mutated forms of presenilins were identified as causative factors in familial Alzheimer's disease. Recent studies show that in addition to Notch and APP, numerous signal transduction pathways are modulated by presenilins, including intracellular calcium signaling. Thus, presenilins appear to have diverse roles. To further understand presenilin function, we searched for Presenilin-interacting genes in Drosophila by performing a genetic modifier screen for enhancers and suppressors of Presenilin-dependent Notch-related phenotypes. We ide... genesis journals http://www.medworm.com/rss/comments.php?id=2213824 Mon, 23 Feb 2009 05:00:00 +0100 2213824 http://www.medworm.com/index.php?rid=2213823&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20487 Local and regulated expression of exogenous genes in the central nervous system is one of the major challenges of modern neuroscience. We have approached this issue by applying the inducible tetracycline system to regulate the expression of EGFP reporter gene in double transgenic rats. We have obtained a strong induction of EGFP only in male testes, which correlated with a high level of rtTA expression only in this organ. To overcome the problem of lack of rtTA protein in the transgenic rat brain, we have delivered this Tet system activator with lentiviral vectors into the dentate gyrus of hippocampus of transgenic EGFP rats. As a result, after systemic application of doxycycline we have obtained inducible, stable and restricted to the desired brain region expression of EGFP. An advantage ... genesis journals http://www.medworm.com/rss/comments.php?id=2213823 Mon, 23 Feb 2009 05:00:00 +0100 2213823 http://www.medworm.com/index.php?rid=2185684&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20454 In this report, we describe the isolation of an otic mesenchyme enhancer in the Brn4 gene. Subsequently, this enhancer was used to drive the expression of Cre recombinase in the otic mesenchyme of transgenic mice. When intercrossed with the ROSA reporter strain, R26R, ß-galactosidase expression is detected in several inner ear structures derived from otic mesenchyme, including the temporal bone, spiral ligament, spiral limbus, and mesenchyme underlying sensory epithelium of the utricle, saccule and semicircular canals. Thus, this Cre pedigree can induce conditional rearrangement of genes in the otic mesenchyme, and will serve as a powerful genetic tool to characterize the function of genes in the mesenchymal tissues of the inner ear. genesis 0:1-5, 2009. © 2009 Wiley-Liss, Inc. (Source: ... genesis journals http://www.medworm.com/rss/comments.php?id=2185684 Fri, 13 Feb 2009 05:00:00 +0100 2185684 http://www.medworm.com/index.php?rid=2176162&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20469 Lentiviral vectors efficiently integrate into the host genome of both dividing and nondividing cells, and so they have been used for stable transgene expression in biological and biomedical studies. However, recent studies have highlighted the risk of insertional mutagenesis and subsequent oncogenesis. Here, we used an integrase-defective lentiviral (IDLV) vector to decrease the chance of random integration and examined the feasibility of lentiviral vector-mediated gene targeting into murine embryonic stem (ES) cells. After transduction with wild-type lentiviral vectors, none of the 512 G418 resistant clones were found to be homologous recombinant clones. Although the transduction efficiency was lower with the IDLV vectors (5.9% of wild-type), successful homologous recombination was observ... genesis journals http://www.medworm.com/rss/comments.php?id=2176162 Wed, 11 Feb 2009 05:00:00 +0100 2176162 http://www.medworm.com/index.php?rid=2176164&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20479 Activation of the bHLH factor Math5 (Atoh7) is an initiating event for mammalian retinal neurogenesis, as it is critically required for retinal ganglion cell formation. However, the cis-regulatory elements and trans-acting factors that control Math5 expression are largely unknown. Using a combination of transgenic mice and bioinformatics, we identified a phylogenetically conserved regulatory element that is required to activate Math5 transcription during early retinal neurogenesis. This element drives retinal expression in vivo, in a cross-species transgenic assay. Previously, Pax6 was shown to be necessary for the initiation of Math5 mRNA expression. We extend this finding by showing that the Math5 retinal enhancer also requires Pax6 for its activation, via Pax6 binding to a highly conser... genesis journals http://www.medworm.com/rss/comments.php?id=2176164 Tue, 10 Feb 2009 05:00:00 +0100 2176164 http://www.medworm.com/index.php?rid=2176163&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20480 We generated a novel mouse model, which expresses the tetracycline-inducible transactivator under the regulation of the endogenous whey acidic protein gene. Using a tet-responsive luciferase reporter transgene, we demonstrated that the Wap-rtTA knockin allele allows a tightly controlled temporal and spatial expression of transgenes in the mammary gland in a ligand-inducible manner. The longitudinal analysis of individual females throughout their reproductive cycles using in vivo bioluminescence imaging confirmed that the expression of the Wap-rtTA knockin allele is highly upregulated during lactation. However, the extent of the transcriptional activation of the targeted Wap locus is dependent on the suckling stimulus and milk retrieval. In addition, we used WAP-rtTA/TetO-H2B-GFP double-tra... genesis journals http://www.medworm.com/rss/comments.php?id=2176163 Tue, 10 Feb 2009 05:00:00 +0100 2176163 http://www.medworm.com/index.php?rid=2158777&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20478 Aph-1 is a multipass transmembrane protein and an essential component of the Presenilin (Psn)-mediated [gamma]-secretase complex. During protease assembly, Aph-1 stabilizes the newly synthesized Psn holoprotein to facilitate generation of the active form of Psn, which is a Psn-NTF/Psn-CTF heterodimer produced through a Presenilinase-initiated endoproteolytic cleavage of the Psn holoprotein. Although it is clear that loss of Aph-1 activity leads to failure of Psn heterodimer formation, little is understood about whether Aph-1 plays a role in regulating [gamma]-secretase activity in addition to assisting Psn maturation. Using various modified Psn forms that do not require endoproteolysis or have a large deletion of the cytosolic loop, we show that in Drosophila Aph-1 is still required for [g... genesis journals http://www.medworm.com/rss/comments.php?id=2158777 Wed, 04 Feb 2009 05:00:00 +0100 2158777 http://www.medworm.com/index.php?rid=2158778&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20475 Presenilin (Psn) is a multipass transmembrane protein that functions as the catalytic subunit of [gamma]-secretase for mediating intramembrane cleavage of type 1 transmembrane proteins. Normally active Psn is in the form of a heterodimer composed by its N-terminal and C-terminal fragments that are generated from a Presenilinase-mediated endoproteolytic cleavage within its large cytosolic loop during assembly of the protease complex. Using the Psn forms that either bypass or disable Presenilinase-mediated endoproteolysis, and a Psn form that has most of the large cytosolic loop deleted, we have established an in vivo system to enable investigations of Psn functional domains in Drosophila. We show that the Presenilinase-mediated endoproteolytic event is not essential for producing Psn activi... genesis journals http://www.medworm.com/rss/comments.php?id=2158778 Tue, 03 Feb 2009 05:00:00 +0100 2158778 http://www.medworm.com/index.php?rid=2123852&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20466 To address the requirement for TGF[beta] signaling in the formation and maintenance of the vascular matrix, we employed lineage-specific mutation of the type II TGF[beta] receptor gene (Tgfbr2) in vascular smooth muscle precursors in mice. In both neural crest- and mesoderm-derived smooth muscle, absence of TGF[beta] receptor function resulted in a poorly organized vascular elastic matrix in late-stage embryos which was prone to dilation and aneurysm. This defect represents a failure to initiate formation of the elastic matrix, rather than a failure to maintain a preexisting matrix. In mutant tissue, lysyl oxidase expression was substantially reduced, which may contribute to the observed pathology. genesis 0:1-7, 2009. © 2009 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2123852 Thu, 22 Jan 2009 05:00:00 +0100 2123852 http://www.medworm.com/index.php?rid=2123851&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20467 The ciliated cells of the node of the mouse embryo contribute to the establishment of left-right patterning via generation of leftward laminar fluid flow and initiation of a left-sided morphogen gradient. Here, we identify FOXJ1CreER2T mice in which expression of Cre recombinase is directed to ciliated node cells. The data demonstrate that foxj1 is expressed specifically in the node throughout the developmental window critical for left-right patterning. In transgenic embryos, Cre expression is detected by immunohistochemistry in ciliated cells of the node. Rosa26R reporter mice, in which expression of lacZ is activated only after Cre-mediated recombination, demonstrate strong and uniform labeling at the node when crossed with FOXJ1CreER2T mice. Cell labeling occurred as early as 0- to 2-so... genesis journals http://www.medworm.com/rss/comments.php?id=2123851 Thu, 22 Jan 2009 05:00:00 +0100 2123851 http://www.medworm.com/index.php?rid=2123850&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20470 The extracellular isoform of superoxide dismutase (EC-SOD, Sod3) plays a protective role against various diseases and injuries mediated by oxidative stress. To investigate the pathophysiological roles of EC-SOD, we generated tetracycline-inducible Sod3 transgenic mice and directed the tissue-specific expression of transgenes by crossing Sod3 transgenic mice with tissue-specific transactivator transgenics. Double transgenic mice with liver-specific expression of Sod3 showed increased EC-SOD levels predominantly in the plasma as the circulating form, whereas double transgenic mice with neuronal-specific expression expressed higher levels of EC-SOD in hippocampus and cortex with intact EC-SOD as the dominant form. EC-SOD protein levels also correlated well with increased SOD activities in dou... genesis journals http://www.medworm.com/rss/comments.php?id=2123850 Thu, 22 Jan 2009 05:00:00 +0100 2123850 http://www.medworm.com/index.php?rid=2123849&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20474 The Cre/lox and FLP/FRT recombination systems have been used extensively for both conditional knockout and cell lineage analysis in mice. Here we report a new multifunctional Cre/FLP dual reporter allele (R26NZG) that exhibits strong and apparently ubiquitous marker expression in embryos and adults. The reporter construct, which is driven by the CAG promoter, was knocked into the ROSA26 locus providing an open chromatin domain for consistent expression and avoiding site-of-integration effects often observed with transgenic reporters. R26NZG directs Cre-dependent nuclear-localized [beta]-galactosidase ([beta]-gal) expression, and can be converted into a Cre-dependent EGFP reporter (R26NG) by germline excision of the FRT-flanked nlslacZ cassette. Alternatively, germline excision of the floxe... genesis journals http://www.medworm.com/rss/comments.php?id=2123849 Thu, 22 Jan 2009 05:00:00 +0100 2123849 http://www.medworm.com/index.php?rid=2093961&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20460 In this study, we characterized the promoter activity of a 1.7 kb sequence in the 5[prime] flanking region of the mouse Deleted in Azoospermia-Like (Dazl) gene. We found the 1.7 kb sequence sufficient to drive robust germ cell-specific expression of green fluorescent protein (GFP) in adult mouse testis and lower levels of expression in adult ovary and in fetal and newborn gonads of both sexes. This expression pattern was confirmed in two independently-derived transgenic mouse lines. In adult testis, Dazl-GFP exhibited a developmentally-regulated, stage-specific expression pattern during spermatogenesis. GFP was highly expressed in spermatocyte stages, with strongest expression in pachytene spermatocytes. Weaker expression was observed in round and elongating spermatids, as well as spermato... genesis journals http://www.medworm.com/rss/comments.php?id=2093961 Fri, 09 Jan 2009 05:00:00 +0100 2093961 http://www.medworm.com/index.php?rid=2069942&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20465 To establish a genetic tool for manipulating the neural stem/progenitor cell (NSC) lineage in a temporally controlled manner, we generated a transgenic mouse line carrying an NSC-specific nestin promoter/enhancer expressing a fusion protein encoding Cre recombinase coupled to modified estrogen receptor ligand-binding domain (ERT2). In the background of the Cre reporter mouse strain Rosa26lacZ, we show that the fusion CreERT2 recombinase is normally silent but can be activated by the estrogen analog tamoxifen both in utero, in infancy, and in adulthood. As assayed by [beta]-galactosidase activity in embryonic stages, tamoxifen activates Cre recombinase exclusively in neurogenic cells and their progeny. This property persists in adult mice, but Cre activity can also be detected in granule ne... genesis journals http://www.medworm.com/rss/comments.php?id=2069942 Thu, 01 Jan 2009 05:00:00 +0100 2069942 http://www.medworm.com/index.php?rid=2069944&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20464 The epidermal growth factor receptor (EGFR) is important for normal homeostasis in a variety of tissues and, when abnormally expressed or mutated, contributes to the development of many diseases. However, in vivo functional studies are hindered by the lack of adult mice lacking EGFR because of the pre- and postnatal lethality of EGFR deficient mice. We generated a conditional allele of Egfr (Egfrtm1Dwt) by flanking exon 3 with loxP sites in order to investigate tissue-specific functions of this widely expressed receptor tyrosine kinase. The activity of the Egfrtm1Dwt allele is indistinguishable from wildtype Egfr. Conversely, the Egfr[Delta] allele, generated by Cre-mediated deletion of exon 3 using the germline EIIa-Cre transgenic line, functions as a null allele. Egfr[Delta]/[Delta] embr... genesis journals http://www.medworm.com/rss/comments.php?id=2069944 Mon, 29 Dec 2008 05:00:00 +0100 2069944 http://www.medworm.com/index.php?rid=2069943&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20473 In this study, we explored the mouse P19 EC cell line as an effective tool to investigate the factors that may play essential roles in mesoderm formation and axial elongation morphogenesis. We first demonstrated that aggregated P19 cells not only exhibited gene expression patterns characteristic of mesoderm formation but also displayed elongation morphogenesis with a distinct anterior-posterior body axis as in the embryo. We then showed by RNA interference that these processes were controlled by various regulators of Wnt signaling pathways, namely [beta]-catenin, Wnt3, Wnt3a, and Wnt5a, in a manner similar to normal embryo development. We further showed by inhibitor treatments that the axial elongation morphogenesis was dependent on the activity of Rho-associated kinase. Because of the con... genesis journals http://www.medworm.com/rss/comments.php?id=2069943 Mon, 29 Dec 2008 05:00:00 +0100 2069943 http://www.medworm.com/index.php?rid=2068655&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20452 Understanding the cellular events that underlie epithelial morphogenesis is a key problem in developmental biology. Here, we describe a new transgenic mouse line that makes it possible to visualize individual cells specifically in the Wolffian duct and ureteric bud, the epithelial structures that give rise to the collecting system of the kidney. myr-Venus, a membrane-associated form of the fluorescent protein Venus, was expressed in the ureteric bud lineage under the control of the Hoxb7 promoter. In Hoxb7/myr-Venus mice, the outlines of all Wolffian duct and ureteric bud epithelial cells are strongly labeled at all stages of urogenital development, allowing the shapes and arrangements of individual cells to be readily observed by confocal microscopy of freshly excised or cultured kidneys.... genesis journals http://www.medworm.com/rss/comments.php?id=2068655 Sun, 28 Dec 2008 05:00:00 +0100 2068655 http://www.medworm.com/index.php?rid=2055151&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20463 We report that some molecularly severe roX1 mutations with no detectable transcript accumulation contribute dramatically to male rescue by autosomal roX1 transgenes. We propose that this represents genetic complementation between a source of roX RNA (the autosomal transgene) and the severely mutated X-linked allele. genesis 0:1-6, 2009. © 2008 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=2055151 Sun, 21 Dec 2008 05:00:00 +0100 2055151 http://www.medworm.com/index.php?rid=2055152&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20459 Aromatase is an enzyme that catalyzes the synthesis of estrogen in gonads and brain. Teleost fish express aromatase (AroB) strongly in the brain facilitating its detailed examination. To understand the function of AroB in the brain, we generated transgenic zebrafish that expresses green fluorescent protein (GFP) driven by the brain aromatase cyp19a1b promoter. GFP was found in the radial glial cells of transgenic larvae and adult fish that overlap with AroB immunoreactivity in the correct temporal and spatial pattern. GFP was also coexpressed with radial cell marker BLBP, but was not in neurons. In addition, GFP expression in the radial glial cells was stimulated by estrogen, same as endogenous AroB expression. Thus, this transgenic line faithfully mimics the regulation of AroB expression ... genesis journals http://www.medworm.com/rss/comments.php?id=2055152 Fri, 19 Dec 2008 05:00:00 +0100 2055152 http://www.medworm.com/index.php?rid=1994670&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20462 We report a transgenic line with highly penetrant cre recombinase activity in the somatotrope cells of the anterior pituitary gland. Expression of the cre transgene is under the control of the locus control region of the human growth hormone gene cluster and the rat growth hormone promoter. Cre recombinase activity was assessed with two different lacZ reporter genes that require excision of a floxed stop sequence for expression: a chick [beta]-actin promoter with the CMV enhancer transgene and a ROSA26 knock-in. Cre activity is detectable in the developing pituitary after initiation of Gh transcription and persists through adulthood with high penetrance in Gh expressing cells and lower penetrance in lactotropes, a cell type that shares a common origin with somatotropes. This Gh-cre transge... genesis journals http://www.medworm.com/rss/comments.php?id=1994670 Fri, 28 Nov 2008 05:00:00 +0100 1994670 http://www.medworm.com/index.php?rid=1994671&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20441 Intracellular membrane trafficking regulates a wide variety of developmental processes, including cell and tissue morphogenesis. Here we report developmental expression of Drosophila Rab11, a small GTP-binding protein, required for both endocytic recycling and exocytosis. Rab11 is expressed in the epithelial cell types of diverse lineages at all developmental stages, beginning from the cellular blastoderm in early embryos to adult primordia and adult tissues, like the columnar epithelia lining male ejaculatory bulb. A robust expression of Rab11 is seen both in the amnioserosa and in the lateral epidermis during embryonic dorsal closure, a morphogenetic event that involves spreading and fusion of the contra-lateral sides of epidermis. Rab11 mutant embryos fail to display the characteristic ... genesis journals http://www.medworm.com/rss/comments.php?id=1994671 Wed, 26 Nov 2008 05:00:00 +0100 1994671 http://www.medworm.com/index.php?rid=1949677&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20449 The aristaless-related gene, Arx, plays a fundamental role in patterning the brain in humans and mice. Arx mutants exhibit lissencephaly among other anomalies. We have cloned a Xenopus aristaless homolog that appears to define specific regions of the developing forebrain. xArx2 is transcribed in blastula through neurula stages, and comes to be restricted to the ventral and lateral telencephalon, lateral diencephalon, neural floor plate of the anterior spinal cord, and somites. In this respect, Arx2 expresses in regions similar to Arx with the exception of the somites. Overexpression enlarges the telencephalon, and interference by means of antisense morpholino-mediated translation knockdown reduces growth of this area. Overexpression and inhibition studies demonstrate that misregulation of ... genesis journals http://www.medworm.com/rss/comments.php?id=1949677 Tue, 11 Nov 2008 05:00:00 +0100 1949677 http://www.medworm.com/index.php?rid=1925934&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20453 Interactions between adjacent epithelial and mesenchymal tissues represent a highly conserved mechanism in embryonic organogenesis. In particular, the ability of the mesenchyme to instruct cellular differentiation of the epithelium is a fundamental requirement for the morphogenesis of tubular structures such as those found in the kidneys, lungs, and the developing male reproductive system. Once the tubular structure has formed, it receives signals from the mesenchyme, which can control proliferation, patterning, and differentiation of the epithelium inside the tube. However, the epithelium is not a "silent partner" in this process, and epithelium-derived factors are often required for proper maintenance of the mesenchymal compartment. Although much emphasis has been placed on the character... genesis journals http://www.medworm.com/rss/comments.php?id=1925934 Sun, 02 Nov 2008 04:00:00 +0100 1925934 http://www.medworm.com/index.php?rid=1919482&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20448 To generate temporally-controlled targeted somatic mutations selectively and efficiently in smooth muscles, we have established a transgenic SMA-Cre-ERT2 mouse line in which the expression of the Tamoxifen-dependent Cre-ERT2 recombinase is under the control of a large genomic DNA segment of the mouse smooth muscle [alpha] actin (SMA) gene, contained in a Bacterial artificial chromosome (Bac). In this transgenic mouse line, Cre-ERT2-mediated recombination of LoxP-flanked target DNA is strictly Tamoxifen-dependent, and efficient in both vascular and visceral smooth muscle cells. Moreover, with the exception of few cardiomyocytes, LoxP-flanked DNA excision is restricted to smooth muscle cells. Thus, SMA-Cre-ERT2 mice should be of great value to analyze gene function in smooth muscles, and to ... genesis journals http://www.medworm.com/rss/comments.php?id=1919482 Fri, 31 Oct 2008 04:00:00 +0100 1919482 http://www.medworm.com/index.php?rid=1919484&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20446 To develop a sensitive and inducible system to study intestinal biology, we generated a transgenic mouse model expressing the reverse tetracycline transactivator rtTA2-M2 under control of the 12.4 kb murine Villin promoter. The newly generated Villin-rtTA2-M2 mice were then bred with the previously developed tetO-HIST1H2BJ/GFP model to assess inducibility and tissue-specificity. Expression of the histone H2B-GFP fusion protein was observed exclusively upon doxycycline induction and was uniformly distributed throughout the intestinal epithelium. The Villin-rtTA2-M2 was also found to drive transgene expression in the developing mouse intestine. Furthermore, we could detect transgene expression in the proximal tubules of the kidney and in a population of alleged gastric progenitor cells. By a... genesis journals http://www.medworm.com/rss/comments.php?id=1919484 Tue, 21 Oct 2008 04:00:00 +0100 1919484 http://www.medworm.com/index.php?rid=1919483&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20447 Pax3-Cre (P3Pro-Cre) transgenic mice have been used for conditional gene deletion and/or lineage tracing in derivatives of neural crest, neural tube, metanephric mesenchyme, and ureteric mesenchyme. However, the extent of its expression in skeletal muscle has not been reported. We investigated the expression of P3Pro-Cre in the skeletal muscle lineage using the R26R reporter and found an unexpected rostrocaudal gradient of expression. By X-gal staining, head, neck, forelimb, diaphragm, and most of the chest wall muscles did not show evidence of Cre expression, whereas all muscle groups posterior of the diaphragm stained blue. Intercostal muscles exhibited a rostrocaudal gradient of staining. The consistency of this expression pattern was demonstrated by using P3Pro-Cre to mutate a conditio... genesis journals http://www.medworm.com/rss/comments.php?id=1919483 Tue, 21 Oct 2008 04:00:00 +0100 1919483 http://www.medworm.com/index.php?rid=1919488&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20416 Chordates originated from a common ancestor(s) shared with two other deuterostome groups, echinoderms and hemichordates, by creating a novel type of tadpole-like larva, which was characterized by a dorsal hollow neural tube and notochord. Recent molecular phylogeny supports the notion that echinoderms and hemichordates form a clade named the Ambulacraria and that, among the chordates, cephalochordates are more basal than urochordates and vertebrates. An aboral-dorsalization hypothesis is proposed to explain how the tadpole-type larva evolved. Embryological comparison of cephalochordates with nonchordate deuterostomes suggests that, because of limited space on the oral side of the ancestral embryo, morphogenesis to form the neural tube and notochord occurred on the aboral side of the embryo... genesis journals http://www.medworm.com/rss/comments.php?id=1919488 Fri, 17 Oct 2008 04:00:00 +0100 1919488 http://www.medworm.com/index.php?rid=1919487&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20438 In female mouse embryos, the paternal X chromosome (Xp) is preferentially inactivated during preimplantation development and trophoblast differentiation. This imprinted X-chromosome inactivation (XCI) is partly due to an activating imprint on the maternal X chromosome (Xm), which is set during oocyte growth. However, the nature of this imprint is unknown. DNA methylation is one candidate, and therefore we examined whether disruptions of the two de novo DNA methyltransferases in growing oocytes affect imprinted XCI. We found that accumulation of histone H3 lysine-27 trimethylation, a hallmark of XCI, occurs normally on the Xp, and not on the Xm, in female blastocysts developed from the mutant oocytes. Furthermore, the allelic expression patterns of X-linked genes including Xist and Tsix wer... genesis journals http://www.medworm.com/rss/comments.php?id=1919487 Fri, 17 Oct 2008 04:00:00 +0100 1919487 http://www.medworm.com/index.php?rid=1919486&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20443 Metamorphosis displays a striking diversity in chordates, a deuterostome phylum that comprises vertebrates, urochordates (tunicates), and cephalochordates (amphioxus). In anuran amphibians, the tadpole loses its tail, develops limbs, and undergoes profound changes at the behavioral, physiological, biochemical, and ecological levels. In ascidian tunicates, the tail is lost and the head tissues are drastically remodeled into the adult animal, whereas in amphioxus, the highly asymmetric larva transforms into a relatively symmetric adult. This wide diversity has led to the proposal that metamorphosis evolved several times independently in the different chordate lineages during evolution. However, the molecular mechanisms involved in metamorphosis are largely unknown outside amphibians and tele... genesis journals http://www.medworm.com/rss/comments.php?id=1919486 Fri, 17 Oct 2008 04:00:00 +0100 1919486 http://www.medworm.com/index.php?rid=1919485&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20439 Single nucleotide polymorphisms (SNPs) may influence protein function possibly contributing to phenotype; yet, for most SNPs their potential influence is unknown. Here, we present a technique in mouse embryonic stem cells that enables high-throughput knock-in (the placement of coding sequences adjacent to a specific endogenous promoter). Our methodology utilizes gene targeting with a combination of two selection cassettes (SA[beta]geo and the HPRT minigene) along with site-specific recombinases (Cre/loxP and FLP/FRT) to efficiently introduce multiple DNA sequences, including enhanced green fluorescent protein (eGFP), adjacent to the DNA topoisomerase 3[beta] (Top3[beta]) promoter. This technology enables rapid and efficient introduction of DNA sequences to a specific location and advances ... genesis journals http://www.medworm.com/rss/comments.php?id=1919485 Fri, 17 Oct 2008 04:00:00 +0100 1919485 http://www.medworm.com/index.php?rid=1887668&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20455 No abstract. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=1887668 Fri, 17 Oct 2008 04:00:00 +0100 1887668 http://www.medworm.com/index.php?rid=1887659&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20458 Top edge: Body wall of the annelid Pristina leidya (Jesse Meik and Ian Swinburne). Right edge: Embryo of the grasshopper, Schistocerca americana (R'ada Massarwa). Bottom edge: Drosophila melanogaster embryo (Courtney Karner). Bottom left corner: Mouse embryo (Marie-Therese Noedl). Left upper row: Adult annelid Pristina leidyi (Michelle Collins and Stephanie Lepage). Middle upper row: Embryo of the squid Loligo pealii (Andrew Chervenak). Right upper row: larva of the colonial ascidian, Botrylloides (Jessica Gray). Left lower row: Chick embryo (Benjamin Schlager). Middle lower row: Xenopus laevis (Brooke Armfield and Zacharias Kontarakis). Right lower row: Third instar CNS and disks of Drosophila melanogaster (Jenna Judge). For more information on the Woods Hole Embrylology Course, visit (So... genesis journals http://www.medworm.com/rss/comments.php?id=1887659 Wed, 01 Oct 2008 04:00:00 +0100 1887659 http://www.medworm.com/index.php?rid=1887667&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20432 The dark-like (dal) mutant mouse has a pleiotropic phenotype that includes dark dorsal hairs and reproductive degeneration. Their pigmentation phenotype is similar to Attractin (Atrn) mutants, which also develop vacuoles throughout the brain. In further characterizing the testicular degeneration of dal mutant males, we found that they had reduced serum testosterone and developed vacuoles in their testes. Genetic crosses placed dal upstream of the melanocortin 1 receptor (Mc1r) and downstream of agouti, although dal suppressed the effect of agouti on pigmentation but not body weight. Atrnmg-3J and dal showed additive effects on pigmentation, testicular vacuolation, and spongiform neurodegeneration, but transgenic overexpression of Attractin-like-1 (Atrnl1), which compensates for loss of ATR... genesis journals http://www.medworm.com/rss/comments.php?id=1887667 Fri, 26 Sep 2008 04:00:00 +0100 1887667 http://www.medworm.com/index.php?rid=1887666&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20434 Eph receptor tyrosine kinases (RTKs) are a highly conserved family of signaling proteins with functions in cellular migration, adhesion, apoptosis, and proliferation during both adult and embryonic life. Here, we describe a knock-in mouse in which EphA1 expression is disrupted via the insertion of an internal ribosome entry site (IRES)-human placental alkaline phosphatase (ALPP) reporter cassette into exon II of the EphA1 gene. This was shown to successfully knockout expression of endogenous EphA1 and enforce expression of the ALPP reporter by the EphA1 promoter. Staining for the ALPP reporter protein demonstrated an epithelially restricted expression pattern in mouse tissues. In EphA1 null mice, two separate phenotypes were identified: abnormal tail development manifesting as a kinky tail... genesis journals http://www.medworm.com/rss/comments.php?id=1887666 Thu, 18 Sep 2008 04:00:00 +0100 1887666 http://www.medworm.com/index.php?rid=1887665&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20433 The cystic fibrosis transmembrane conductance regulator (CFTR) gene encodes a cAMP-regulated chloride channel that is important in controlling the exchange of fluid and electrolytes across epithelial cells. Mutation of CFTR can lead to cystic fibrosis (CF), the most common lethal genetic disease in Caucasians. CF is a systemic illness with multiple organ systems affected including pulmonary, gastrointestinal, pancreatic, immune, endocrine, and reproductive systems. To understand the role of CFTR in the various tissues in which it is expressed, we generated a murine conditional null allele of Cftr (Cftrfl10) in which loxP sites were inserted around exon 10 of the Cftr gene. The Cftrfl10 allele was validated by generating constitutive Cftr null (Cftr[Delta]10) mice using the protamine-cre sy... genesis journals http://www.medworm.com/rss/comments.php?id=1887665 Thu, 18 Sep 2008 04:00:00 +0100 1887665 http://www.medworm.com/index.php?rid=1887664&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20424 Developing neural tubes are bilaterally symmetric in all vertebrate embryos, irrespective of the presence of gene networks that generate left-right asymmetry. To explore the mechanisms that underlie the bilaterally symmetric formation of the neural tube, we examined a medaka (Oryzias latipes) dominant mutant, Oot, the neural tube of which transiently lacks normal symmetry in the optic tectum. We found that spatial changes in isthmic fgf8 expression do not occur on one side of the mutant, resulting in a transient desynchronized expression that correlates with tectal asymmetry. The application of exogenous FGF8 on one side of a wild-type embryo mimics the Oot phenotype, indicating that the bilaterally equivalent expression of isthmic fgf8 is crucial for the bilaterally symmetric development ... genesis journals http://www.medworm.com/rss/comments.php?id=1887664 Thu, 18 Sep 2008 04:00:00 +0100 1887664 http://www.medworm.com/index.php?rid=1887663&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20426 Taking advantage of a mutant estrogen receptor ligand binding domain (ERT2), we developed novel Caspase fusion proteins for inducible apoptosis. We show that Caspase-ERT2 fusion proteins become specifically activated by the synthetic ligand 4-OH- tamoxifen and rapidly induce apoptotic cell death in human, murine, and zebrafish cells. This novel tool for targeted cell ablation greatly facilitates the generation of disease models as well as developmental and regeneration studies in model organisms. genesis 46:530-536, 2008. © 2008 Wiley-Liss, Inc. (Source: genesis) genesis journals http://www.medworm.com/rss/comments.php?id=1887663 Thu, 18 Sep 2008 04:00:00 +0100 1887663 http://www.medworm.com/index.php?rid=1887662&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20427 Wnt signaling plays an important role in regulating cortical and hippocampal development, but many of the other molecular mechanisms underlying dorsal telencephalic development are largely unknown. We are taking advantage of the highly regionalized expression patterns of signaling components of the Wnt pathway to generate new mouse lines that will be useful for studying forebrain development. Here, we describe a transgenic mouse line where Cre is driven by the promoter of the Wnt receptor, Frizzled10. In these mice, Cre activity is mainly detected in the dorsal telencephalon during development and is confined to the pyramidal cell fields in the adult hippocampus. The Cre recombinase has very high efficiency when assayed by crossing the transgenic line with the ROSA26 reporter line. Thus, t... genesis journals http://www.medworm.com/rss/comments.php?id=1887662 Thu, 18 Sep 2008 04:00:00 +0100 1887662 http://www.medworm.com/index.php?rid=1887660&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20398 This report introduces a gonadotrope-specific cre transgenic mouse capable of ablating floxed genes in mature pituitary gonadotropes. Initial analysis of this transgenic line, Tg(Lhb-cre)1Sac, reveals that expression is limited to the pituitary cells that produce luteinizing hormone beta, beginning appropriately at e17.5. Cre activity is detectable by a reporter gene in nearly every LH[beta]-producing cell, but the remaining hormone-producing cell types and other organs exhibit little to no activity. We used the Tg(Lhb-cre)1Sac strain to assess the role Pitx2 in gonadotrope function. The gonadotrope-specific Pitx2 knockout mice exhibit normal expression of LH[beta], sexual maturation, and fertility, suggesting that Pitx2 is not required for gonadotrope maintenance or for regulated producti... genesis journals http://www.medworm.com/rss/comments.php?id=1887660 Thu, 18 Sep 2008 04:00:00 +0100 1887660 http://www.medworm.com/index.php?rid=1887661&cid=s_33772_50_f&fid=33772&url=http%3A%2F%2Fdx.doi.org%2F10.1002%252Fdvg.20410 Foxa2 is a forkhead transcription factor expressed in the node, notochord, floorplate, and definitive endoderm and is required in the foregut endoderm for the normal development of the endoderm-derived organs, such as the liver, lung and pancreas. To conditionally inactivate genes in these tissues and organs, we have targeted a Cre recombinase into Exon 1 of the Foxa2 gene. We show, upon crossing to the ROSA26 reporter mice, that Cre expression from the Foxa2iCre knock-in allele specifically activates [beta]-galactosidase expression in the node, notochord, floorplate, and endoderm. In addition, we detect Cre recombination activity in the endoderm-derived organs including lung, liver, pancreas, and gastrointestinal tract throughout development. These results demonstrate that the Foxa2iCre k... genesis journals http://www.medworm.com/rss/comments.php?id=1887661 Tue, 16 Sep 2008 04:00:00 +0100 1887661