pinecone/core-2020-05-10-deduplication huggingface dataset:
A Novel Transgenic Technique That Allows Specific Marking of the Neural Crest Cell Lineage in Mice

AbstractNeural crest cells are embryonic, multipotent stem cells that give rise to various cell/tissue types and thus serve as a good model system for the study of cell specification and mechanisms of cell differentiation. For analysis of neural crest cell lineage, an efficient method has been devised for manipulating the mouse genome through the Cre–loxP system. We generated transgenic mice harboring a Cre gene driven by a promoter of protein 0 (P0). To detect the Cre-mediated DNA recombination, we crossed P0-Cre transgenic mice with CAG-CAT-Z indicator transgenic mice. The CAG-CAT-Z Tg line carries a lacZ gene downstream of a chicken β-actin promoter and a “stuffer” fragment flanked by two loxP sequences, so that lacZ is expressed only when the stuffer is removed by the action of Cre recombinase. In three different P0-Cre lines crossed with CAG-CAT-Z Tg, embryos carrying both transgenes showed lacZ expression in tissues derived from neural crest cells, such as spinal dorsal root ganglia, sympathetic nervous system, enteric nervous system, and ventral craniofacial mesenchyme at stages later than 9.0 dpc. These findings give some insights into neural crest cell differentiation in mammals. We believe that P0-Cre transgenic mice will facilitate many interesting experiments, including lineage analysis, purification, and genetic manipulation of the mammalian neural crest cells
Discrete Enhancer Elements Mediate Selective Responsiveness of Enhancer of split Complex Genes to Common Transcriptional Activators

AbstractIn Drosophila, genes of the Enhancer of split Complex [E(spl)-C] are important components of the Notch (N) cell–cell signaling pathway, which is utilized in imaginal discs to effect a series of cell fate decisions during adult peripheral nervous system development. Seven genes in the complex encode basic helix-loop-helix (bHLH) transcriptional repressors, while 4 others encode members of the Bearded family of small proteins. A striking diversity is observed in the imaginal disc expression patterns of the various E(spl)-C genes, suggestive of a diversity of function, but the mechanistic basis of this variety has not been elucidated. Here we present strong evidence from promoter–reporter transgene experiments that regulation at the transcriptional level is primarily responsible. Certain E(spl)-C genes were known previously to be direct targets of transcriptional activation both by the N-signal-dependent activator Suppressor of Hairless [Su(H)] and by the proneural bHLH proteins achaete and scute. Our extensive sequence analysis of the promoter-proximal upstream regions of 12 transcription units in the E(spl)-C reveals that such dual transcriptional activation is likely to be the rule for at least 10 of the 12 genes. We next show that the very different wing imaginal disc expression patterns of E(spl)m4 and E(spl)mγ are a property of small (200–300 bp), evolutionarily conserved transcriptional enhancer elements, which can confer these distinct patterns on a heterologous promoter despite their considerable structural similarity [each having three Su(H) and two proneural protein binding sites]. We also demonstrate that the characteristic inactivity of the E(spl)mγ enhancer in the notum and margin territories of the wing disc can be overcome by elevated activity of the N receptor. We conclude that the distinctive expression patterns of E(spl)-C genes in imaginal tissues depend to a significant degree on the capacity of their transcriptional cis-regulatory apparatus to respond selectively to direct proneural- and Su(H)-mediated activation, often in only a subset of the territories and cells in which these modes of regulation are operative
The TGF-β2 Isoform Is Both a Required and Sufficient Inducer of Murine Hair Follicle Morphogenesis

AbstractHair follicle development serves as an excellent model to study control of organ morphogenesis. Three specific isoforms of TGF-β exist which exhibit a distinct pattern of expression during hair follicle morphogenesis. To clarify the still elusive role of these factors in hair follicle development, we have used a combined genetic and functional approach: analysis of hair follicle development in mice with disruptions of the TGF-β1, 2, and 3 genes was coupled with a direct functional test of the effect of added purified factors on fetal hair follicle development in skin organ cultures. TGF-β2 null mice exhibited a profound delay of hair follicle morphogenesis, with a 50% reduced number of hair follicles. In contrast to hair follicle development, growth and differentiation of interfollicular keratinocytes proceeded unimpaired. Unlike TGF-β2−/− mice, mice with a disruption of the TGF-β1 gene showed slightly advanced hair follicle formation, while lack of the TGF-β3 gene did not have any effects. Treatment of wild-type, embryonic skin explants (E14.5 or E15.5) with TGF-β2 protein in either soluble form or slow release beads induced hair follicle development and epidermal hyperplasia, while similar TGF-β1 treatment exerted suppressive effects. Thus, the TGF-β2 isoform plays a specific role, not shared by the other TGF-β isoforms, as an inducer of hair follicle morphogenesis and is both required and sufficient to promote this process
Characterization of GDF-10 Expression Patterns and Null Mice

AbstractGrowth/differentiation factor-10 (GDF-10) is a TGF-β family member highly related to bone morphogenetic protein-3. In order to determine the biological function of GDF-10, we carried out a detailed analysis of the expression pattern of GDF-10 and characterized GDF-10-null mice that we generated by gene targeting. During embryogenesis GDF-10 is expressed prominently in developing skeletal structures both in the craniofacial region and in the vertebral column. In adult animals, GDF-10 is expressed at high levels in the brain, where GDF-10 is localized primarily to cells in the Purkinje cell layer of the cerebellum, and in the uterus, where the expression levels of GDF-10 are regulated both during the menstrual cycle and during pregnancy. Despite the high levels of GDF-10 expression in these tissues, we found no obvious abnormalities in GDF-10-knockout mice with respect to the development of these tissues. These findings suggest either that GDF-10 plays no regulatory role in these tissues or that its function is redundant with that of other growth factor-like molecules
Neurofibromin Deficiency in Mice Causes Exencephaly and Is a Modifier for Splotch Neural Tube Defects

AbstractNeural tube defects are common and serious human congenital anomalies. These malformations have a multifactorial etiology and can be reproduced in mouse models by mutations of numerous individual genes and by perturbation of multiple environmental factors. The identification of specific genetic interactions affecting neural tube closure will facilitate our understanding of molecular pathways regulating normal neural development and will enhance our ability to predict and modify the incidence of spina bifida and other neural tube defects. Here, we report a genetic interaction between Nf1, encoding the intracellular signal transduction protein neurofibromin, and Pax3, a transcription factor gene mutated in the Splotch mouse. Both Pax3 and Nf1 are important for the development of neural crest-derived structures and the central nervous system. Splotch is an established model of folate-sensitive neural tube defects, and homozygous mutant embryos develop spina bifida and sometimes exencephaly. Neural development is grossly normal in heterozygotes and neural tube defects are not seen. In contrast, we found a low incidence of neural tube defects in heterozygous Splotch mice that also harbored a mutation in one Nf1 allele. All compound homozygotes had severe neural tube defects and died earlier in embryogenesis than either Nf1−/− or Sp−/− embryos. We also report occasional exencephaly in Nf1−/− mice and identify more subtle CNS abnormalities in normal-appearing Nf1−/− embryos. Though other genetic loci and environmental factors affect the incidence of neural tube defects in Splotch mice, these results establish Nf1 as the first known gene to act as a modifier of neural tube defects in Splotch
Ectopic Expression of the nude Gene Induces Hyperproliferation and Defects in Differentiation: Implications for the Self-Renewal of Cutaneous Epithelia

AbstractNude mice are characterized by the absence of visible hair, epidermal defects, and the failure to develop a thymus. This phenotype results from loss-of-function mutations in Whn (Hfh11), a winged-helix transcription factor. In murine epidermis and hair follicles, endogenous whn expression is induced as epithelial cells initiate terminal differentiation. Using the promoter for the differentiation marker involucrin, transgenic mice that ectopically express whn in stratified squamous epithelia, hair follicles, and the transitional epithelium of the urinary tract were generated. Transgenic epidermis and hair follicles displayed impaired terminal differentiation and a subset of hair defects, such as delayed growth, a waved coat, and curly whiskers, correlated with decreased transforming growth factor (TGF)-α expression. The exogenous Whn protein also stimulated epithelial cell multiplication. In the epidermis, basal keratinocytes exhibited hyperproliferation, though transgene expression was restricted to suprabasal, postmitotic cells. Hair follicles failed to enter telogen (a resting period) and remained continuously in an abnormal anagen (the growth phase of the hair cycle). Ureter epithelium developed severe hyperplasia, leading to the obstruction of urine outflow and death from hydronephrosis. Though an immune infiltrate was present occasionally in transgenic skin, the infiltrate was not the primary cause of the epithelial hyperproliferation, as the immune reaction was not observed in all affected transgenics, and the transgene induced identical skin and urinary tract abnormalities in immunodeficient Rag1-null mice. Given the effects of the transgene on cell proliferation and TGFα expression, the results suggest that Whn modulates growth factor production by differentiating epithelial cells, thereby regulating the balance between proliferative and postmitotic populations in self-renewing epithelia
Impaired Mammary Gland Development in Cyl-1−/− Mice during Pregnancy and Lactation Is Epithelial Cell Autonomous

AbstractA specific defect of mice lacking cyclin D1 (Cyl-1−/−) is impaired development of the mammary gland during pregnancy. Here we show that when tissue from Cyl-1−/− mammary gland was transplanted into empty mammary fat pad of wild-type mice, the abnormal phenotype was maintained, indicating that it is epithelial cell autonomous. Nevertheless, in pregnancy the early proliferative response, which is characterized by extensive side branching, still occurs in the absence of cyclin D1. However, the response is atypical due to a marked reduction in the formation of accompanying alveoli. This reduction and delay in alveolar development persists throughout pregnancy. Moreover, although prolactin synthesis and release appear to be normal, lactogenesis is severely compromised. Consistent with the appearance of numerous side branches, progesterone receptor expression was readily detected in the mammary tissue of pregnant Cyl-1−/− mice, although there was a significant change in the ratio of the two (A and B) receptor isoforms. In Cyl-1−/− mammary glands during late pregnancy there was a decrease in the abundance of total and phosphorylated Stat5a, as well as delayed onset and substantial diminution of milk protein expression. The biochemical analysis suggests that there is a cumulative delay in growth and differentiation of the mammary gland during pregnancy that results in a severely compromised gland when, at parturition, further development is curtailed by the abrupt change in hormonal milieu
Interference with Brachyury Function Inhibits Convergent Extension, Causes Apoptosis, and Reveals Separate Requirements in the FGF and Activin Signalling Pathways

AbstractBrachyury plays a key role in mesoderm formation during vertebrate development. Absence of the gene results in loss of posterior mesoderm and failure of the notochord to differentiate, while misexpression of Brachyury in the prospective ectoderm of Xenopus results in ectopic mesoderm formation. Brachyury is therefore both necessary and sufficient for posterior mesoderm formation. Here we present a detailed cellular and molecular analysis of the consequences of inhibiting Brachyury function during Xenopus development. Our results show that Brachyury is required for the convergent extension movements of gastrulation, for mesoderm differentiation in response to FGF, and for the survival of posterior mesodermal cells in both Xenopus and mouse
Cloning of Rat Fibrillin-2 cDNA and Its Role in Branching Morphogenesis of Embryonic Lung

AbstractFibrillin-2 is an extracellular matrix protein. It is associated with elastic fibers in several tissues and is believed to serve as a ligand for αvβ3 integrin, the latter being a known morphogen. In this study, the role of fibrillin-2 in lung development was investigated. Also, rat fibrillin-2 cDNA was isolated and sequenced and its spatiotemporal expression determined. It had ∼88% homology with human fibrillin-2 and had Ca2+ binding epidermal growth factor-like domains, transforming growth factor-β binding protein motifs, and two RGD binding sites. Northern blot analysis revealed an ∼10-kb transcript, and fibrillin-2 expression was developmentally regulated, and it paralleled that of tropoelastin. At day 13 of gestation, fibrillin-2 was expressed in the mesenchyme and at the epithelial:mesenchymal interface. From day 13 to 19 of gestation, its expression intensified and was confined around the tracheobronchial airways, while it lessened during the postnatal period. Immunoprecipitation revealed an ∼350-kDa band by SDS–PAGE. Treatment with fibrillin-2 antisense oligodeoxynucleotide induced dysmorphogenesis of the lung explants. They were smaller and had rudimentary lung bud branches, collapsed conducting airways, and loose expanded mesenchyme. Concomitantly, fibrillin-2 mRNA, antibody reactivity in the explants, and fibrillin-2-specific radioincorporation were reduced. Anti-αv and -laminin antibody reactivity and their respective incorporated specific radioactivities were unaltered. These data indicate that fibrillin-2 modulates organogenesis of the lung in the context of epithelial:mesenchymal interactions. Conceivably, the collapse of the conducting airways may also be related to the perturbed biology of the fibrillin-2 interacting protein, i.e., elastin, the latter being critical for the normal biophysiology of the lungs
A Paraxial Exclusion Zone Creates Patterned Cranial Neural Crest Cell Outgrowth Adjacent to Rhombomeres 3 and 5

AbstractCranial neural crest cell migration is patterned, with neural crest cell-free zones adjacent to rhombomere (R) 3 and R5. These zones have been suggested to result from death of premigratory neural crest cells via upregulation of BMP-4 and Msx-2 in R3 and R5, consequent to R2-, R4-, and R6-derived signals. We reinvestigated this model and found that cell death detected by acridine orange staining in avian embryos varied widely numerically and in pattern, but with a tendency for an elevated zone centered at the R2/3 boundary. In situ hybridization of BMP-4 mRNA resolved to centers at R3 and R5 but Msx-2 resolved to the R2/3 border with only a faint smear from R5 to R6. Outgrowth of neural crest cells was less in isolated R3 cultures than in R1+2, R2, and R4 cultures, but R3 showed neither a decrease in outgrowth of neural crest cells nor an increase in cell death when cocultured with R1+2, R2, or R4. In addition, in serum-free culture, exogenous BMP-4 strikingly reduced neural crest cell outgrowth from R1+2 and R4 as well as R3. Thus we cannot confirm the role of intraneural cell death in patterning rhombomeric neural crest outgrowth. However, grafting quail R2 or R4 adjacent to the chick hindbrain demonstrated a neural crest cell exclusion zone next to R3 and R5. We suggest that one important pattern determinant for rhombomeric neural crest cell migration involves the microenvironment next to the neural tube
posterior end mark 3 (pem-3), an Ascidian Maternally Expressed Gene with Localized mRNA Encodes a Protein with Caenorhabditis elegans MEX-3-like KH Domains

AbstractMaternal factors localized in the posterior-vegetal cytoplasm of an ascidian egg are essential for cell specification and pattern formation of the embryo. The molecular identification of the localized factors and the elucidation of the machinery associated with the localization are therefore key research subjects. I report here the isolation and characterization of a novel maternally expressed gene, posterior end mark 3 (pem-3). The pem-3 cDNA was obtained from a cDNA library of fertilized egg mRNAs subtracted with gastrula mRNAs of Ciona savignyi. As in the case of pem (Yoshida et al., 1996, Development 122, 2005–2012), the pem-3 maternal transcript was gradually concentrated after fertilization in the posterior-vegetal cytoplasm of the egg, and it later marked the posterior end of developing embryos. The PEM-3 protein was also detected in the posterior end of early embryos. The nucleotide sequence predicted that pem-3 encodes a probable RNA-binding protein with two KH domains that have an extensive similarity with those of Caenorhabditis elegans MEX-3. MEX-3 is also localized in nematode embryos (Draper et al., 1996, Cell 87, 205–216), suggesting that PEM-3 is a candidate homologue of MEX-3. In addition to maternal expression, a zygotic transcript of pem-3 and its gene product were detected in cells of the neural plate, mesenchyme, and epidermis of embryos after the neural-plate stage. Inhibition of zygotic expression using an antisense oligonucleotide resulted in the development of abnormal larvae without sensory pigment cells, suggesting that the zygotic PEM-3 plays a role in the differentiation of the brain of the ascidian larva
CnOtx, a Member of the Otx Gene Family, Has a Role in Cell Movement in Hydra

AbstractOtx genes have been identified in a variety of organisms and are commonly associated with the patterning of anterior structures. In some vertebrates, Otx genes are also expressed in the prechordal mesoderm, where they may have a role in cell movement. Here we report the characterization of CnOtx, an Otx gene in hydra, thereby providing evidence that Otx genes appeared early in metazoan evolution. CnOtx is expressed at high levels in developing buds and aggregates, where it appears to have a role in the cell movements that are involved in the formation of new axes. Further, the gene is expressed at a low level throughout the body column of hydra. This latter pattern may reflect a role for CnOtx in specifying tissue as competent to be anterior, although the gene does not have a direct role in the formation of the head
Slug Is a Mediator of Epithelial–Mesenchymal Cell Transformation in the Developing Chicken Heart

AbstractAn epithelial–mesenchymal cell transformation occurs during the development of the endocardial cushions in the atrioventricular (AV) canal of the heart. We hypothesized that the transcription factor Slug is required for this epithelial–mesenchymal cell transformation since Slug is required for similar transformations during gastrulation and neural crest differentiation in chicken embryos. We found by RT-PCR and immunostaining that the temporal and spatial localization of Slug in the embryonic chicken heart is consistent with a role for Slug in endocardial cushion formation. Moreover, we found that Slug expression by AV canal endothelial cells is induced by a signal provided by AV canal myocardium. Slug appears to be required for epithelial–mesenchymal cell transformation in the chicken heart since treatment of AV canal explants with antisense Slug oligodeoxynucleotides inhibited mesenchymal cell formation in vitro. Antisense Slug oligodeoxynucleotides prevented endothelial cell–cell separation, suggesting that Slug acts early in the transformation pathway
Negative Regulation of Dorsoventral Signaling by the Homeotic Gene Ultrabithorax during Haltere Development in Drosophila

AbstractGrowth and patterning during Drosophila wing development are mediated by signaling from its dorsoventral (D/V) organizer. In the metathorax, wing development is essentially suppressed by the homeotic selector gene Ultrabithorax (Ubx) to mediate development of a pair of tiny balancing organs, the halteres. Here we show that expression of Ubx in the haltere D/V boundary down-regulates its D/V organizer signaling compared to that of the wing D/V boundary. Somatic loss of Ubx from the haltere D/V boundary thus results in the formation of a wing-type D/V organizer in the haltere field. Long-distance signaling from this organizer was analyzed by assaying the ability of a Ubx− clone induced in the haltere D/V boundary to effect homeotic transformation of capitellum cells away from the boundary. The clonally restored wing D/V organizer in mosaic halteres not only enhanced the homeotic transformation of Ubx− cells in the capitellum but also caused homeotic transformation of even Ubx+ cells in a genetic background known to induce excessive cell proliferation in the imaginal discs. In addition to demonstrating a non-cell-autonomous role for Ubx during haltere development, these results reveal distinct spatial roles of Ubx during maintenance of cell fate and patterning in the halteres
Activation of a UBC4-Dependent Pathway of Ubiquitin Conjugation during Postnatal Development of the Rat Testis

AbstractDuring spermatogenesis, germ cells undergo mitotic and meiotic divisions to form haploid round spermatids which mature to functional elongated spermatozoa. During this process there occurs remodeling of cell structure and loss of most of the cytoplasm and a large fraction of cellular proteins. To evaluate the role of the ubiquitin proteolytic system in this protein loss, we measured levels of ubiquitinated proteins and rates of ubiquitin conjugation in extracts of testes from rats of different ages. Endogenous ubiquitin–protein conjugates increased till day 30 and then reached a plateau. In parallel, there was a progressive increase in the rate of conjugation of ubiquitin to proteins in testis extracts from these animals. To test the importance of two major ubiquitin conjugating enzyme families in the conjugation, immunoprecipitation of UBC2 or UBC4 from 10- and 30-day-old testis extracts was carried out and the remaining conjugation activity in supernatants was assayed. Depletion of either enzyme family resulted in decreased conjugation. However, most of the conjugation activity and, more importantly, the increased conjugation during development were UBC4-dependent. Immunocytochemistry demonstrated a marked increase in expression of UBC4 in spermatids, consistent with the UBC4-dependent activation of conjugation seen in vitro. In situ hybridization studies evaluated the contribution of various UBC4 isoforms to this induction. UBC4-1 mRNA was expressed in most cells. UBC4-2 mRNA was restricted to germ cells with high levels of expression in round and elongated spermatids. UBC4-testis had previously been shown to be expressed only in spermatids. Our data suggest that induction of various UBC4 isoforms activates overall conjugation and plays an important role in the cellular remodeling and protein loss occurring during spermatogenesis