pinecone/core-2020-05-10-deduplication huggingface dataset:
Microtubule Dependence of Chromosome Cycles inXenopus laevisBlastomeres under the Influence of a DNA Synthesis Inhibitor, Aphidicolin

AbstractThe spindle-assembly checkpoint of the cell cycle develops inXenopus laevisembryos at the midblastula transition (MBT). Our previous experiments using animal-cap blastomeres indicate that the checkpoint is regulated by a mechanism that depends on age, but not on the nucleocytoplasmic (N/C) ratio (Clute and Masui, 1995). In the present study, the time of appearance of the spindle-assembly checkpoint is examined in animal-cap blastomeres whose N/C ratio is reduced by treatment with aphidicolin. Animal-cap blastomeres treated with aphidicolin from the 2-cell stage cleave more slowly after 4th cleavage, in a dose-dependent manner, but cleavage and chromosome cycles continue up to the 11th to 13th cleavage and then arrest. Blastomeres treated with aphidicolin have a reduced DNA content and N/C ratio compared to control blastomeres of the same age. Nevertheless, nocodazole-sensitive chromosome cycles appear at the same time as in control blastomeres, at 3 to 5 hr after 5th cleavage, regardless of the N/C ratio. The arrest in interphase caused by treating blastula stage animal caps with aphidicolin can be reversed by treatment with caffeine. The caffeine-induced mitosis becomes sensitive to nocodazole after the MBT, but not before. Therefore, the same mechanism which stabilizes maturation-promoting factor activity in the absence of a mitotic spindle also operates after the MBT in blastomeres that are treated with aphidicolin, if mitosis is induced by caffeine. This mechanism may involve the translation of a maternal mRNA at the time of the MBT, as suggested previously
In VitroPreselection of Gene-Trapped Embryonic Stem Cell Clones for Characterizing Novel Developmentally Regulated Genes in the Mouse

AbstractWe have developed anin vitrogene trap screen for novel murine genes that allows one to determine, prior to making chimeric or transgenic animals, if these genes are expressed in one or more specific embryonic tissues. Totipotent embryonic stem (ES) cells are infected with a retroviral gene trap construct encoding a selectablelacZ/neoRfusion gene, which is expressed only if the gene trap inserts within an active transcription unit. G418-resistant ES cell clones are induced to differentiatein vitro,and neurons, glia, myocytes, and chondrocytes are screened for expression of β-galactosidase (β-gal). cDNAs of the gene trap transcripts are obtained by 5′ rapid amplification of cDNA ends and are sequenced to determine if they represent novel genes.In situhybridization analyses show that trapped genes are expressedin vivowithin the cell types that express β-galin vitro.Gene traps and their wild-type alleles are characterized in terms of copy number, alternate splicing of their transcripts, and the proportion of endogenous mRNA sequence that is replaced bylacZ/neoRin the hybrid gene trap transcript. This approach, which we term “in vitropreselection,” is more economical than standardin vivogene trap screening because tissue-specific expression of probable knockout alleles is verified before transgenic animals are generated. These results also highlight the utility of ES cell differentiationin vitroas a method with which to study the molecular mechanisms regulating the specification and commitment of a variety of cell and tissue types
N-cadherin Promotes the Commitment and Differentiation of Skeletal Muscle Precursor Cells

AbstractCells with the potential to form skeletal muscle are present in the chick embryo prior to gastrulation. Muscle differentiation begins after gastrulation within the somites. The role of cadherin-mediated adhesion in the commitment and differentiation of skeletal muscle precursor cells was examined by analyzing the expression of cell–cell adhesion molecules in cultures of epiblast, segmental plate, and somite cells and by determining the effects of adhesion-perturbing antibodies on the accumulation of MyoD and sarcomeric myosin. Cultured primitive streak stage epiblast cells downregulate E-cadherin and upregulate N-cadherin. This switch in cadherin expression also occursin vivoas epiblast cells enter the primitive streak. Although MyoD protein is present in cells with N- or E-cadherin, only cells with N-cadherin differentiate into skeletal muscle. In contrast to the primitive streak stage epiblast cells, prestreak epiblast cells maintain the expression of E-cadherinin vitro.While the majority of prestreak cells contain MyoD, only a few synthesize myosin. Treatment of primitive streak stage epiblast cells with function-perturbing antibodies to N-cadherin resulted in an inhibition of myosin accumulation and a decrease in the percentage of cells with MyoD. Segmental plate and somite cells are similar to primitive streak stage epiblast cells in that most differentiated into skeletal muscle when cultured in serum-free medium. While function-perturbing antibodies to N-cadherin inhibited the accumulation of myosin in these mesoderm cells, the number of MyoD positive cells was unaffected in somite cultures and only partially reduced in segmental plate cultures. These results suggest that N-cadherin-mediated cell–cell adhesion is involved in both the commitment of muscle precursors and their terminal differentiation
In VivoRegulation of Cytostatic Activity inXenopusMetaphase II-Arrested Oocytes

AbstractMetaphase II arrest ofXenopusoocyte is characterized by the presence of M-phase-promoting factor (MPF) and of a microtubular spindle, both of which are stable in the presence of protein synthesis inhibitors. We studiedin vivothis equilibrium state that is settled during meiotic maturation. At time of germinal vesicle breakdown (GVBD), cdc2 kinase and MAP kinase activities are stimulated. A component of the cyclin ubiquitin ligase, CDC27, is phosphorylated at the same time and remains phosphorylated until fertilization, indicating that an important component of the ligase complex is modified as early as GVBD. During a first period extending from GVBD until the cortical anchorage of the metaphase II spindle, homogeneous pools of cdc2 kinase and mitogen-activated protein (MAP) kinase activities are present in oocyte and are strictly dependent on protein turnover, since protein synthesis inhibition induces their total inactivation and drives oocytes into interphase. The metaphase II spindle, once anchored into the cortex, is no more sensitive to protein synthesis inhibition, likewise MAP kinase activity. During this cellular arrest, cdc2 kinase is divided into two distinctly regulated pools. The first one contains cyclin B that actively turns over and is subjected to a microtubular checkpoint. The second one is stable. Alteration of intracellular compartmentation of metaphase II oocytes either by gentle centrifugation or by cold shock inactivates MAP kinase and targets all cyclin B molecules for full destruction. We therefore suggest that MAP kinase participates to the cytostatic activity by preventing part of cyclin B molecules from entering the ubiquitination/degradation machinery which is still turned on in metaphase II oocytes
TheCaenorhabditis elegansOrphan Nuclear Hormone Receptor Genenhr-2Functions in Early Embryonic Development

AbstractWe have identified aCaenorhabditis elegansgene,nhr-2,that is a member of the nuclear hormone receptor superfamily of transcription factors and defines a new subclass of the superfamily.nhr-2messenger RNA is expressed in the maternal germline and during the first half of embryogenesis. Zygotic expression ofnhr-2begins by the 16-cell stage, making it one of the earliest genes known to be transcribed in the embryo. Immunolocalization detects NHR-2 protein in embryonic nuclei as early as the 2-cell stage. The protein is present in every nucleus until the 16- to 20-cell stage. Subsequently, expression continues in many, but not all, cell lineages, becoming progressively restricted to the anterior and dorsal regions of the embryo and disappearing during the initial stages of morphogenesis. Disruption ofnhr-2function with antisense RNA results in embryonic and early larval arrest, indicating that the gene has an essential function in embryonic development.nhr-2does not correspond to known mutations mapped to the same genetic interval, and will provide an entry point for further study of a heretofore uncharacterized zygotic gene regulatory pathway
Differential Expression of Two γ-Tubulin Isoforms during Gametogenesis and Development inDrosophila

AbstractPrevious work identified a γ-tubulin gene, γTub23C,inDrosophila(Zhenget al.,1991). We now report identification of a second γ-tubulin gene, γTub37CD.Immunoblot analysis and immunolocalization show that γTub37CD and γTub23C are differentially expressed during gametogenesis and development. During oogenesis, γTub23C was detected at centrosomes and in the cytoplasm of mitotic germ cells, but was not detected in germ cells following completion of mitosis. Conversely, γTub37CD was not detected in proliferating germ cells, but appeared to accumulate in germ cells during egg chamber development. Neither γ-tubulin isoform was detected at the anterior or posterior poles of developing oocytes. During spermatogenesis, only γTub23C was detected at centrosomes, where it showed cell cycle- and differentiation-dependent organization. During the transition into the first meiotic division, γTub23C became organized as a corpuscular focus at centrioles until completion of meiosis II. During postmeiotic spermatid differentiation, γTub23C was detected first as a rod and then as a collar-like structure near the juncture of the nucleus and the elongating flagellum, but was not detected in bundles of mature sperm. The germline-specific CDC25 encoded bytwineis required for organization of γTub23C into corpuscular focus in spermatocytes, but not for separation of centriole pairs in M-phase or postmeiotic organization of γTub23C at centrioles. Following reconstitution of a canonical centrosome at fertilization, only γTub37CD was detected at centrosomes in syncytial embryos, but both γTub37CD and γTub23C were detected at centrosomes in cellularized embryos. Colocalization of these two isoforms suggests that γTub23C and γTub37CD both contain structural features of γ-tubulins essential for localization to centrosomes
Delta-1 Is a Regulator of Neurogenesis in the Vertebrate Retina

AbstractIn the retina, cell fate determination is thought to be regulated by a series of local cell–cell interactions. Evidence suggests that retinal precursors utilize Notch-mediated intercellular signaling to regulate their fates. However, the identity of the endogenous ligand and its role in the Notch-signaling pathway is not well understood. We have identified C-Delta-1 as the putative endogenous ligand for Notch, in the developing chick retina.C-Delta-1is coexpressed spatially and temporally withC-Notch-1and their expression is associated with the temporal aspects of cell birth in the developing retina. This suggests that Delta–Notch signaling is utilized to maintain progenitors in an uncommitted state and that a subtle fluctuation in this signaling helps to sort out competent cells during successive cell-fate determination. We have tested the latter possibility in the specification of the ganglion cells. In early stages of retinal development when ganglion cells are the predominant cells born, decreasingC-Delta-1expression with antisense oligonucleotides increases the proportion of RA4 antigen-expressing ganglion cells which are recruited predominantly in the periphery. Conversely, use of exogenousDrosophilaDelta leads to a decrease in the RA4 antigen-expressing ganglion cells. Our results suggest that C-Delta-1 activation of the Notch pathway regulates the specification of retinal neurons in general and of ganglion cells in particular
Division of Labor of Schwann Cell Integrins during Migration on Peripheral Nerve Extracellular Matrix Ligands

AbstractMyelination of the peripheral nervous system (PNS) requires the migration of Schwann cells during both development and regeneration. We have characterized the expression pattern of Schwann cell integrins and analyzed their role in migration on different ECM substrates known to be present within the PNS. We found that Schwann cells in cell culture express four β1 integrins, α1β1, α2β1, α6β1, and another unidentified β1 integrin, as well as two αv integrins, αvβ3 and αvβ8. Using the Varani migration assay, we found that laminin-1, laminin-2 (merosin), and fibronectin increased Schwann cell migration, while vitronectin and collagen did not increase migration compared to an uncoated plastic substrate. Schwann cell migration on laminin-1 and laminin-2 (merosin) was blocked by antibodies against β1 integrins, but not affected by RGD peptides or antibodies against β3 integrins. In contrast, migration on fibronectin was unaffected by antibodies against β1 and β3 integrins but was blocked by RGD peptides. Thisin vitrostudy shows that there is a division of labor of Schwann cell integrins in the regulation of migration on peripheral nerve ECM components; β1 integrins mediate migration on laminin-1 and laminin-2 (merosin), while αv integrins mediate migration on fibronectin. Taken together, these results suggest that multiple interactions between Schwann cell integrins and ECM within the PNS will contribute to Schwann cell migration during myelination of the PNS
Smad5 Induces Ventral Fates inXenopusEmbryo

AbstractThe Smad proteins have been implicated in the intracellular signaling of transforming growth factor-β (TGF-β) ligands. Here we describe the function of Smad5 in earlyXenopusdevelopment. Misexpression of Smad5 in the embryo causes ventralization and induces ventral mesoderm. Moreover, Smad5 induces epidermis in dissociated ectoderm cells which would otherwise form neural tissue. Both of these activities require Smad4 (DPC4) activity. We propose that Smad5 acts downstream of the BMP4 signaling pathway inXenopusembryos and directs the formation of ventral mesoderm and epidermis
Cytoplasmically Anchored Plakoglobin Induces a WNT-like Phenotype inXenopus

AbstractPlakoglobin is one of two vertebrate proteins closely related to theDrosophilasegment polarity gene product armadillo. Overexpression of plakoglobin induces neural axis duplication inXenopusand the exogenous plakoglobin is localized to nuclei (Karnovsky, A., and Klymkowsky, M.W.,Proc. Natl. Acad. Sci. USA92, 4255, 1995; Rubenstein, A.,et al., Dev. Genet.,1997, in press). We have carried out a series of experiments to test whether the nuclear localization of plakoglobin is required for its inductive effects. Prior to the midblastula transition exogenous plakoglobin is cytoplasmic and concentrated in the cortical regions of blastomeres; after the midblastula transition exogenous plakoglobin accumulates in embryonic nuclei. The addition of a “nuclear localization sequence” does not change the timing of plakoglobin's nuclear localization, suggesting that it is anchored in the cytoplasm prior to the midblastula transition. Next, we constructed two “membrane-anchored” forms of plakoglobin. These are exclusively cytoplasmic; yet both were as effective at producing a “Wnt-like” axis duplication as were “free,” unfettered forms of plakoglobin. Moreover, expression of anchored plakoglobins had no apparent effect on the cytoplasmic or nuclear levels of β-catenin. These data indicate that plakoglobin can act cytoplasmically to generate a WNT-like phenotype. Taken together with the ventralizing effects of a mutant form of the XTcf-3 transcription factor, described by Molenaaret al. Cell86, 391, 1996, we speculate that in the earlyXenopusembryo, activation of plakoglobin (or β-catenin) inhibits the activity of XTcf-3 or a XTcf-3-like factor
Analysis of Neural Crest Cell Migration in Splotch Mice Using a Neural Crest-Specific LacZ Reporter

AbstractStudies on the mouseSplotch(Sp) mutation, a deletion in the transcription factor Pax-3, have revealed that Pax-3 is essential for normal development of the neural crest. We have investigated the defect in neural crest development using a Wnt-l::LacZ reporter construct to mark neural crest cells. Staining embryos for β-galactosidase activity at different developmental stages revealed a severe reduction in the number of neural crest cells which emigrated from the neural tube at the vagal and rostral trunk levels. At the caudal thoracic, lumbar, and sacral levels there was a complete loss of neural crest cell emigration. In contrast to previous work in culture, we saw no evidence for any delay in the onset of neural crest cell migration at anterior levels. Pax-3 is expressed in the dorsal neural tube, where the neural crest cells originate, in migrating neural crest cells, and in somitic cells along the migratory pathway. Hence, it is not clear which aspect of the Pax-3 expression accounts for the observed phenotype. We addressed this problem by transplanting neural tissue between mouse and chick embryos. Our studies indicate that the defect in theSplotchmutation is not intrinsic to the neural crest cells themselves, but appears to reflect inappropriate cell interactions either within the neural tube or between the neural tube and the somite
Activation of the Wnt Signaling Pathway: A Molecular Mechanism for Lithium Action

AbstractGlycogen synthase kinase-3β (GSK-3β/zeste-white-3/shaggy) is a negative regulator of the wnt signaling pathway which plays a central role in the development of invertebrates and vertebrates; loss of function and dominant negative mutations in GSK-3β lead to activation of the wnt pathway inDrosophilaandXenopus.We now provide evidence that lithium activates downstream components of the wnt signaling pathwayin vivo,leading to accumulation of β-catenin protein. Our data indicate that this activation of the wnt pathway is a consequence of inhibition of GSK-3β by lithium. Using a novel assay for GSK-3β in oocytes, we show that lithium inhibits GSK-3β from species as diverse asDictyostelium discoideumandXenopus laevis,providing a biochemical mechanism for the action of lithium on the development of these organisms. Lithium treatment also leads to activation of an AP-1–luciferase reporter inXenopusembryos, consistent with previous observations that GSK-3β inhibits c-jun activity. Activation of the wnt pathway with a dominant negative form of GSK-3β is inhibited by myo-inositol, similar to the previously described effect of coinjecting myo-inositol with lithium. The mechanism by which myo-inositol inhibits both dominant negative GSK-3β and lithium remains uncertain
TheSTUDGene Is Required for Male-Specific Cytokinesis after Telophase II of Meiosis inArabidopsis thaliana

AbstractDuring male meiosis in wild-typeArabidopsisthe pollen mother cell (PMC) undergoes two meiotic nuclear divisions in the absence of cell division. Only after telophase II is a wall formed which partitions the PMC into four microspores. Each microspore undergoes two subsequent mitotic divisions to produce one vegetative cell and two sperm cells in the mature pollen grain. In this paper we describe the isolation and the phenotypic characterization of mutations in theSTUD(STD) gene, which is specifically required for male-specific cytokinesis after telophase II of meiosis. Although the male meiotic nuclear divisions are normal instdmutant plants, no walls are formed resulting in a tetranucleate microspore. Despite the absence of cell division in the PMC, postmeiotic development in the coenocytic microspore proceeds relatively normally, resulting in the formation of large pollen grains which contain four vegetative nuclei and up to eight sperm cells. Interestingly, these enlarged pollen grains which contain multiple vegetative nuclei and extra sperm cells behave as single male gametophytes, producing only single pollen tubes and resulting in partial male fertility instdmutant plants. Characterization of the process of pollen development and pollen function instdmutants thus reveals two different types of developmental regulation. Each of the four nuclei found in astdmicrospore following meiosis is capable of independently undergoing the complete mitotic cell division (including cytokinesis) which the single nucleus of a wild-type microspore would normally undertake. The ability of the four meiotic products to independently continue through mitosis does not depend on their division into separate cells, but is controlled by some subcellular component found within the coenocytic micropsore. By contrast, the maturestdpollen grain functions as a unit and produces only a single pollen tube despite the presence of multiple nuclei within the vegetative cell, suggesting that this process is controlled at the cellular level independently of the extra subcellular components
Distinct Signal/Response Mechanisms Regulatepax1andQmyoDActivation in Sclerotomal and Myotomal Lineages of Quail Somites

AbstractPax1andQmyoDare early sclerotome and myotome-specific genes that are activated in epithelial somites of quail embryos in response to axial notochord/neural tube signals.In situhybridization experiments reveal that the developmental kinetics of activation ofpax1andQmyoDdiffer greatly, suggesting that myotome and sclerotome specification are controlled by distinct developmental mechanisms.pax1activation always occurs in somite IV throughout development, indicating thatpax1regulation is tightly coordinated with early steps in somite maturation. In contrast,QmyoDis delayed and does not occur until embryos have 12–14 somites. At this time,QmyoDis the first of the myogenic regulatory factor (MRF) genes to be activated in preexisting somites in a rapid, anterior to posterior progression until the 22 somite stage, after which timeQmyoDis activated in somite I immediately following somite formation. Experiments involving transplantation of newly formed somites to ectopic sites along the anterior to posterior embryonic axis were performed to distinguish the contributions of axial signals and somite response pathways to the developmental regulation ofpax1andQmyoD.These studies show thatpax1activation is regulated by somite formation and maturation, not by the availability of axial signals, which are expressed prior to somite formation. In contrast, the delayed activation ofQmyoDis controlled by developmental regulation of the production of axial signals as well as by the competence of somites to respond to these signals. These somite transplantation studies, therefore, provide a basis for understanding the different developmental kinetics of activation ofpax1andQmyoDduring sclerotome and myotome specification, and suggest specific molecular models for the developmental regulation of myotome and sclerotome formation in somites through distinct signal/response pathways
Role of the Dlx Homeobox Genes in Proximodistal Patterning of the Branchial Arches: Mutations of Dlx-1, Dlx-2, and Dlx-1 and -2 Alter Morphogenesis of Proximal Skeletal and Soft Tissue Structures Derived from the First and Second Arches

AbstractThe Dlx homeobox gene family is expressed in a complex pattern within the embryonic craniofacial ectoderm and ectomesenchyme. A previous study established that Dlx-2 is essential for development of proximal regions of the murine first and second branchial arches. Here we describe the craniofacial phenotype of mice with mutations in Dlx-1 and Dlx-1 and -2. The skeletal and soft tissue analyses of mice with Dlx-1 and Dlx-1 and -2 mutations provide additional evidence that the Dlx genes regulate proximodistal patterning of the branchial arches. This analysis also elucidates distinct and overlapping roles for Dlx-1 and Dlx-2 in craniofacial development. Furthermore, mice lacking both Dlx-1 and -2 have unique abnormalities, including the absence of maxillary molars. Dlx-1 and -2 are expressed in the proximal and distal first and second arches, yet only the proximal regions are abnormal. The nested expression patterns of Dlx-1, -2, -3, -5, and -6 provide evidence for a model that predicts the region-specific requirements for each gene. Finally, the Dlx-2 and Dlx-1 and -2 mutants have ectopic skull components that resemble bones and cartilages found in phylogenetically more primitive vertebrates