pinecone/core-2020-05-10-deduplication huggingface dataset:
ADrosophilaKinesin-like Protein, Klp38B, Functions during Meiosis, Mitosis, and Segmentation

AbstractWe show thatklp38B,isolated as a mutation that dominantly prolongs blastoderm mitotic cycles in Drosophila, encodes a Drosophila kinesin-like protein. Further genetic analyses show that Klp38B not only functions during mitosis, but is also required for meiosis and abdominal segmentation. Sequence comparisons suggest that Klp38B encodes an amino-terminal microtubule motor domain, a central α-helical coiled-coil domain, and a C-terminal globular domain. Evidence that Klp38B is required during meiosis is that flies transheterozygous for mutations in bothklp38Bandnodhave a high frequency of 4th chromosome meiotic nondisjunction. Nod is a chromokinesin, a chromosome binding kinesin, that is believed to provide astral-exclusion forces during the metaphase stage of meiosis. Evidence that Klp38B is required during mitosis is that embryos from female germline clones ofklp38Bmutations have holes in the cuticle similar to a zygoticstring(dCDC25) phenotype. Also, anti-Klp38B antibody injection into precellularization blastoderm embryos causes developmental arrest and the formation of circular mitotic figures. We speculate, based on these phenotypes, that Klp38B is a chromokinesin that provides astral-exclusion forces on the chromosomes during meiosis and mitosis. Consistent with this hypothesis, we have identified an HMG-1 homologous region on Klp38B that could potentially bind AT-rich DNA sequences. Finally, we show thatklp38Bmutations have defects in abdominal segmentation, suggesting that Klp38B, like Xenopus chromokinesin Xklp1, might be involved in polar granule formation
The Secretory Protein Sec8 Is Required for Paraxial Mesoderm Formation in the Mouse

AbstractThesec8gene, isolated in a gene trap screen in embryonic stem cells, is required for paraxial mesoderm formation in the mouse. Homozygoussec8mutant embryos initiate gastrulation but are unable to progress beyond the primitive streak stage and die shortly afterward. The genomic locus and cDNA of thesec8gene have been cloned. An open reading frame in the cDNA encodes a 971-amino-acid leucine-rich protein, similar to rat rSec8. A description of the mutant phenotype and the cloning of the gene is presented here and the results are considered in light of the possibility that the Sec8 protein is involved in secretion
The POU Geneceh-18Promotes Gonadal Sheath Cell Differentiation and Function Required for Meiotic Maturation and Ovulation inCaenorhabditis elegans

AbstractInCaenorhabditis elegans,specialized contractile myoepithelial cells of the somatic gonad, the gonadal sheath cells, are closely apposed to oocytes and are required for normal meiotic maturation and ovulation. Previously we found that mutations in theceh-18gene, which encodes a POU-class homeoprotein expressed in sheath cells, result in oocyte defects. To determine the basis for these oocyte defects, we have used time-lapse video Nomarski microscopy to observe meiotic maturation, ovulation, and early embryogenesis inceh-18mutants. Inceh-18mutants sheath cell contractions are weaker, less frequent, and uncoordinated throughout the sequence of ovulation events, and ovulation is defective. Defective ovulation can result in the formation of endomitotic oocytes in the gonad, the formation of haploid embryos, and reversals in embryonic polarity.ceh-18mutant oocytes exhibit defects prior to nuclear envelope breakdown, suggesting that they are physiologically different from the wild type. We observed delays in meiotic maturation, as well as maturation out of the normal spatial and temporal sequence, suggesting that proximal sheath cells directly or indirectly promote and spatially restrict meiotic maturation. Analysis of sheath cell differentiation inceh-18mutants using antibodies to proteins of the contractile apparatus reveals that although contractile proteins are expressed, the sheath cells appear disorganized. Transmission electron microscopy reveals thatceh-18mutant sheath cells are morphologically irregular and only loosely cover oocytes. Taken together, these observations indicate thatceh-18is a crucial determinant of sheath cell differentiation, a function required for normal meiotic maturation and ovulation
The Fucose Sulfate Polymer of Egg Jelly Binds to Sperm REJ and Is the Inducer of the Sea Urchin Sperm Acrosome Reaction

AbstractThe sea urchin sperm cell is an advantageous model for studying ligand-mediated exocytosis. Sperm can be obtained in vast quantities and induced to undergo exocytosis of the acrosomal vesicle with great synchrony. During sea urchin fertilization, egg jelly (EJ) triggers the sperm acrosome reaction (AR) which is required for sperm binding and fusion with the egg. Uncertainty exists as to the exact biochemical nature of the AR inducer. The following study was performed in an attempt to clarify the nature of the inducer. EJ from individual females (Strongylocentrotus purpuratus) was analyzed on SDS–PAGE gels. Each female had a unique composition of EJ macromolecules, but all females possessed the previously described fucose sulfate polymer (FSP). Two electrophoretic isotypes of FSP were discovered; 87% of females had only one isotype and 13% had both. Both FSP isotypes bound to the REJ protein (receptor for egg jelly) purified from sperm. The two FSP isotypes had almost equal potency in inducing the AR. EJ was fractionated by DEAE chromatography in 6 M urea/4% β-mercaptoethanol. All AR-inducing activity coeluted with FSP. FSP, purified by trypsin digestion followed by dialysis, was twice as active as the non-trypsin-digested control at inducing the AR. EJ was digested with proteinase K, boiled in detergent and β-mercaptoethanol, and subjected to sucrose density gradient sedimentation. The FSP and AR activity had superimposable sedimentation patterns. Purified FSP had no associated peptide component. Sperm from individual males differed in the concentration dependency of purified FSP to induce the AR. The data indicate that the 138/82 kDa EJ glycoproteins, previously thought to act as AR inducers, do not appear to be involved in triggering the AR. The data are consistent with the hypothesis that FSP is the only inducer of the AR of this sea urchin species
posterior end mark 2 (pem-2),pem-4,pem-5, andpem-6: Maternal Genes with Localized mRNA in the Ascidian Embryo

AbstractThe posterior–vegetal cytoplasm of an ascidian egg contains maternal factors required for pattern formation and cell specification of the embryo. We report here the isolation and characterization of cDNA clones for novel maternal genes,posterior end mark 2(pem-2),pem-4,pem-5, andpem-6. We obtained these clones from a cDNA library ofCiona savignyifertilized egg mRNAs subtracted with gastrula mRNAs by examining the localization of the corresponding mRNAs of randomly selected clones by whole-mountin situhybridization. As in the case ofpem, all of these mRNAs were localized in the posterior–vegetal cytoplasm of the egg, and they later marked the posterior end of early embryos. The predicted amino acid sequence suggested that PEM-2 contains a signal for nuclear localization, an src homology 3 (SH3) domain, and a consensus sequence of the CDC24 family guanine nucleotide dissociation stimulators (GDSs). PEM-4 has a signal for nuclear localization and three C2H2-type zinc finger motifs, while PEM-5 and PEM-6 show no similarity to known proteins. These results provide further evidence that the ascidian egg contains maternal messages that are localized in the posterior–vegetal cytoplasm
Isolation and Characterization of Chondroitin Sulfate Proteoglycans from Embryonic Quail That Influence Neural Crest Cell Behavior

AbstractThe movement of neural crest cells is controlled in part by extracellular matrix. Aggrecan, the chondroitin sulfate proteoglycan from adult cartilage, curtails the ability of neural crest cells to adhere, spread, and move across otherwise favorable matrix substratesin vitro.Our aim was to isolate, characterize, and compare the structure and effect on neural crest cells of aggrecan and proteoglycans purified from the tissues through which neural crest cells migrate. We metabolically radiolabeled proteoglycans in E2.5 quail embryos and isolated and characterized proteoglycans from E3.3 quail trunk and limb bud. The major labeled proteoglycan was highly negatively charged, similar in hydrodynamic size to chick limb bud versican/PG-M, smaller than adult cartilage aggrecan but larger than reported for embryonic sternal cartilage aggrecan. The molecular weight of the iodinated core protein was about 400 kDa, which is more than reported for aggrecan but less than that of chick versican/PG-M. The proteoglycan bore chondroitin sulfate glycosaminoglycan chains of 45 kDa, which is larger than those of aggrecan. It lacked dermatan sulfate, heparan sulfate, or keratan sulfate chains. It bound to collagen type I, like aggrecan, but not to fibronectin (unlike versican/PG-M), collagen type IV, or laminin-1 in solid-phase assays and it bound to hyaluronate in gel-shift assays. When added at concentrations between 10 and 30 μg/ml to substrates of fibronectin, trunk proteoglycan inhibited neural crest cell spreading and migration. Attenuation of cell spreading was shown to be the most sensitive and titratable measure of the effect on neural crest cells. This effect was sensitive to digestion with chondroitinase ABC. Similar cell behavior was also produced by aggrecan and the small dermatan sulfate proteoglycan decorin; however, 30-fold more aggrecan was required to produce an effect of similar magnitude. When added in solution to neural crest cells which were already spread and migrating on fibronectin, the embryonic proteoglycan rapidly and reversibly caused complete rounding of the cells, being at least 30-fold more potent than aggrecan in this activity
Altered Forebrain and Hindbrain Development in Mice Mutant for theGsh-2Homeobox Gene

AbstractThe patterning of the mammalian brain is orchestrated by a large battery of regulatory genes. Here we examine the developmental function of theGsh-2nonclustered homeobox gene. Whole-mount and serial sectionin situhybridizations have been used to better defineGsh-2expression domains within the developing forebrain, midbrain, and hindbrain.Gsh-2transcripts are shown to be particularly abundant in the hindbrain and within the developing ganglionic eminences of the forebrain. In addition, mice carrying a targeted mutation ofGsh-2have been generated and characterized. Homozygous mutants uniformly failed to survive more than 1 day following birth. At the physiologic level the mutants experienced apnea and reduced levels of hemoglobin oxygenation. Histologically, the mutant brains had striking alterations of discrete components. In the forebrain the lateral ganglionic eminence was reduced in size. In the hindbrain, the area postrema, an important cardiorespiratory chemosensory center, was absent. The contiguous nucleus tractus solitarius, involved in integrating sensory input to maintain homeostasis, was also severely malformed in mutants. Immunohistochemistry was used to examine the mutant brains for alterations in the distribution of markers specific for serotonergic and cholinergic neurons. In addition,in situhybridizations were used to define expression patterns of the Dlx 2 and Nkx 2.1 homeobox genes in Gsh-2 mutant mice. The mutant lateral ganglionic eminences showed an abnormal absence of Dlx 2 expression. These results better define the genetic program of development of the mammalian brain, support neuromeric models of brain development, and further suggest similar patterning function for homeobox genes in phylogenetically diverse organisms
WNT-7a Induces Axonal Remodeling and Increases Synapsin I Levels in Cerebellar Neurons

AbstractWNT factors play a key role in early patterning of the embryo. However, expression ofWntgenes after cell commitment suggests additional roles in later developmental processes. We report here thatWnt-7ais expressed in cerebellar granule cell neurons as they begin to extend processes and form synapses. WNT-7a increases axonal spreading and branching in cultured granule cells. Moreover, WNT-7a increases the levels of synapsin I, a presynaptic protein involved in synapse formation and function. Lithium mimics WNT-7a in granule cells by inhibiting GSK-3β, a component of the WNT signaling pathway. These results suggest a direct effect of WNT-7a in the regulation of neuronal cytoskeleton and synapsin I in granule cell neurons. We propose that WNT proteins have a novel function in the formation of neuronal connections
Mesoderm Is Required for the Formation of a Segmented Endodermal Cell Layer in the LeechHelobdella

AbstractThe homeobox geneLox3is expressed in a segmentally iterated pattern within the endoderm of the leechHelobdella.We use that expression here to study endoderm differentiation following experimental ablations of mesoderm.Lox3RNA was first detected byin situhybridization at the stage when a definitive cellular endoderm is formed from its syncytial precursor and was never observed in derivatives of other germ layers. Expression is initially distributed throughout the endoderm, but rapidly disappears from specific regions of the nascent gut wall so as to produce a pattern of segmental stripes. The stripe pattern differs markedly between midgut organs, with thin stripes ofLox3expression in the intercaecal constrictions of the crop and wide stripes ofLox3expression marking the caecal bulges of the intestine.Lox3expression in the rectum is not obviously segmental. Ablation of segmental mesoderm in the earlyHelobdellaembryo prevents the formation of definitive endoderm and the expression ofLox3RNA and leads to abnormalities in the morphogenesis of the gut tube. These endodermal deficits are precisely coextensive with the zone of mesodermal deficiency, suggesting that the mesoderm normally acts to promote the formation of the endodermal cell layer via local cell interactions. The segmental pattern ofLox3expression is largely unaffected in portions of the endoderm surrounding such deficits, suggesting that endodermal segmentation is not established by lateral interactions within that tissue layer. Rather, we propose that the segmental organization of the endoderm is imprinted by vertical interactions with the segmental mesoderm
A Critical Period of Ear Development Controlled by Distinct Populations of Ciliated Cells in the Zebrafish

AbstractThe zebrafish (Danio rerio) is a useful model system for analyzing development of the inner ear. A number of mutations affecting the inner ear have been identified. Here we investigate the initial stages of otolith morphogenesis in wild-type embryos as well as inmonolith(mnl) mutant embryos, which fail to form anterior otoliths but otherwise appear normal. Otolith growth is initiated at 18–18.5 h by localized accretion of free-moving precursor particles. This process, referred to as otolith seeding, is regulated by two classes of cilia: First, kinocilia of precociously forming hair cells (tether cells) bind seeding particles, thereby localizing otolith formation. Tether cells usually occur in pairs at the anterior and posterior ends of the ear. Despite the presence of functional kinocilia, tether cells initially appear immature and do not acquire the characteristics of mature hair cells until ∼21.5 h. Second, beating cilia distributed throughout the ear agitate seeding particles, thereby inhibiting premature agglutination. Constraining particles with laser tweezers caused them to fuse into large untethered masses. Bringing such masses into contact with tethered otoliths caused them to fuse, greatly enhancing otolith growth. Selectively enhancing one otolith greatly inhibited growth of the second, creating an imbalance that persisted for many days. Seeding particles and beating cilia disappear soon after 24 h, and the rate of otolith growth decreases by nearly 90%. Inmnlmutant embryos, tethers and beating cilia are distributed normally, but anterior otoliths fail to form in 80–85% of mutant ears. The binding properties of seeding particles appear normal, as shown by their ability to fuse when entrapped by laser tweezers and their binding to posterior tethers. We infer that anterior tethers have a weakened ability to bind seeding particles inmnlembryos. Immobilizingmnlembryos with the anterior end of the ear oriented downward effectively concentrated the dense seeding particles near the anterior tethers and permitted all to form anterior otoliths. However, immobilizingmnlembryos after 24 h when seeding particles were depleted did not facilitate anterior otolith formation. Together, these data demonstrate that the ability to initiate otolith formation is limited to a critical period, from 18.5 to 24 h, and that interfering with the functions of tether cell kinocilia or beating cilia impairs otolith seeding and subsequent otolith morphogenesis
The Homeobox GenePV.1Mediates Specification of the Prospective Neural Ectoderm inXenopusEmbryos

AbstractBone morphogenetic protein 4 (BMP4), a member of the TGF β superfamily, has been implicated in the dorsoventral specification of both mesoderm and ectoderm. High levels of BMP4 signaling appear to specify ventral lineages, while lower levels are causally associated with the development of dorsal lineages. We have previously identified a homeobox-containing transcription factor (PV.1) which is a likely mediator of the ventralizing effects of BMP4 in the mesoderm. Here we provide evidence thatPV.1also functions downstream of BMP4 in the patterning of ectoderm, specifying epidermal and suppressing neural gene expression.PV.1is expressed in the prospective neuroectoderm at the time of ectodermal fate determination. BMP4 andxSmad1(a downstream effector of BMP4) inducePV.1in uncommitted ectoderm and the dominant negative form of the BMP4 receptor (DN-BR) blocksPV.1expression. In animal pole explantsPV.1counteracts the neuralizing effects of chordin and the DN-BR and restores them to their original epidermal fate. To address the physiological significance of these observations we employed an animal cap transplantation system and demonstrated that overexpression ofPV.1in the prospective neuroectoderm specifically blocks neurogenesis in intact embryos. Thus,PV.1plays an important role in the ventralization of both mesoderm and ectoderm. We have previously shown thatPV.1is also preferentially expressed in the ventral endoderm, suggesting that this transcription factor may be involved in the ventralization of all three germ layers
Activation of the Receptor Tyrosine Kinase Kit Is Required for the Proliferation of Melanoblasts in the Mouse Embryo

AbstractThe development of neural crest-derived melanocytes, as well as haematopoietic and germ cells, is affected by mutations of theKitandMgfgenes, which lead to dominant spotting (W) or steel (Sl) phenotypes.Mgfcodes for the ligand of the receptor tyrosine kinase encoded by theKitlocus.KitW-v, a point mutation exerting a dominant negative effect, causes a substantial reduction in tyrosine kinase activity of the Kit receptor and leads to a characteristic pigmentation phenotype, namely dilute coat colour and a white ventral and head spot with reduced pigmentation of the feet and tail in the heterozygous animal, as well as slight anaemia. Homozygous animals lack coat pigmentation and are severely anaemic and infertile. Dct is a marker for cells of the melanoblast lineage. In order to study these cells in detail we have generated transgenic mouse lines carrying thelacZreporter under the control of theDctpromoter and have used the embryonic expression of the reporter to identify early melanoblasts before they begin to produce pigment. Our transgenic lines have simplified the study of melanoblasts in the mouse embryo, and by crossing our mice withKitW-vmutants we have been able to identify the midgestation stages at which melanoblasts rely critically on Mgf/Kit interactions. We conclude that the survival of immature melanoblasts depends crucially upon Kit signalling up until E11, and later in development Kit plays a vital role in melanoblast proliferation. Our data do not describe a dependence upon Kit for melanoblast migration or differentiation
Characterization of the Role of Cadherin in Regulating Cell Adhesion during Sea Urchin Development

AbstractDuring development, the modulation of cadherin adhesive function is proposed to control various morphogenetic events including epithelial–mesenchymal conversions and tubulogenesis, although the mechanisms responsible for regulating cadherin activity during these events remain unclear. In order to gain insights into the regulation of cadherin function during morphogenesis, we utilized the sea urchin embryo as a model system to study the regulation of cadherin localization during epithelial–mesenchymal conversion and convergent–extension movements. Polyclonal antibodies raised against the cytoplasmic domain of a cloned sea urchin cadherin recognize three major polypeptides ofMr320, 140, and 125 kDa and specifically stain adherens junctions, and to a lesser extent, lateral membrane domains in all epithelial tissues of the embryo. Analysis of embryos during gastrulation demonstrates that changes in cadherin localization are observed in cells undergoing an epithelial–mesenchymal conversion. Ingression of primary mesenchyme cells is accompanied by the rapid loss of junctional cadherin staining and the coincident accumulation of cadherin in intracellular organelles. These data are consistent with the idea that the deadhesion of mesenchymal cells from neighboring epithelial cells involves the regulated endocytosis of cell surface cadherin molecules. Conversely, neither cadherin abundance nor localization is altered in cells of the gut which undergo convergent–extension movements during the formation of the archenteron. This observation indicates that these movements do not require the loss of junctional cadherin molecules. Instead, the necessary balance between adhesion and motility may be achieved by regulating the expression of different subtypes of cadherin molecules or modifying interactions between cadherins and catenins, proteins that bind the cytoplasmic domain of cadherin and are necessary for cadherin adhesive function. To address cadherin function at the molecular level, we used a partial cDNA representing the conserved cytoplasmic domain to identify a novel cadherin molecule in the sea urchinLytechinus variegatus.The deduced amino acid sequence of LvG-cadherin (for Goliath-cadherin) predicts that it is a transmembrane protein with an apparent relative molecular mass of 303 kDa. The cytoplasmic domain shows significant sequence identity to that of vertebrate classic cadherins. However, the extracellular domain is distinguished from its vertebrate counterparts by both an increased number of cadherin-specific repeats and the presence of four EGF-like repeats proximal to the transmembrane domain. Taken together, these data are consistent with the hypothesis that the sea urchin possesses several cadherins, including a novel member of the cadherin family, and that the dynamic regulation of cadherin localization plays a role in epithelial to mesenchymal conversions during gastrulation