pinecone/core-2020-05-10-deduplication huggingface dataset:
Specific Glycoconjugates Are Present at the Oolemma of the Fertilization Site in the Egg ofDiscoglossus pictus(Anurans) and Bind Spermatozoa in anin VitroAssay

AbstractIn the egg of the anuranDiscoglossus pictus,the site of fertilization is restricted to the central portion of an animal hemisphere indentation (thedimple). Previous studies showed that the acrosome reaction ofD. pictussperm is triggered in the jelly, and yet sperm arrive at the dimple surface with the plasma membrane at an early stage of vesiculation. Reactivity of the dimple surface with specific lectins suggests that fucose might be utilized as a marker of glycoproteins located at the dimple surface. In this paper, proteins of the egg surface were labeled with the membrane impermeable sulfo-NHS-biotin. Four main bands of 200, 230, 260, and 270 kDa labeled only at the dimple surface, although they were detected in the cortex of the whole egg. The 270-kDa band reacted withGalanthus nivalisagglutinin only in the cortex of the dimple, suggesting that this band is differently glycosylated according to its localization. The α-l-fucose-specific lectinUlex europaeusagglutinin I was utilized both in lectin blotting and in affinity chromatography and cross-reacted with the 200- and 270/260-kDa bands. Furthermore, two polypeptides were obtained by exposure of intact eggs to lysylendoproteinase C. They were also reactive toUlex europaeusagglutinin I. The 200- and 270/260-kDa bands were eluted from the acrylamide gels and adsorbed to polystyrene beads. An assay for sperm binding to 200-kDa glycoprotein-bound beads was developed. Sperm stuck to the beads before but not after Ca-ionophore treatment. When the beads were coated with the 270/260-kDa glycoproteins, binding occurred after ionophore treatment. In these assays, the 200- and 270/260-kDa glycoproteins competitively inhibited sperm binding to the beads coated with the corresponding glycoprotein. These results indicate that the assayed glycoproteins, located either in the glycocalyx or in the plasma membrane of the fertilization site, are involved in sperm binding
The Cell Lineage of a Polyclad Turbellarian Embryo Reveals Close Similarity to Coelomate Spiralians

AbstractRecent molecular evidence suggests the turbellarian Platyhelminthes may represent the extant basal members of the Spiralia and therefore probably exhibit ancient features of the spiralian developmental program. The stereotypic quartet spiral cleavage pattern of the polyclad turbellarian embryo, among other features, indicates that this group may be closely related to the ancestral flatworm; however, polyclad embryos have been the subject of few experimental studies. Here we report the results of a cell lineage analysis of the embryo of the polycladHoploplana inquilinabased on microinjection of DiI into cleavage-stage blastomeres following formation of each of the four quartets of micromeres. The first quartet gives rise to most of the lateral and anterior ectoderm of the Müller's larva; the second quartet forms largely dorsal and ventral ectoderm as well as the circular muscles; the third quartet forms only small clones of ectoderm; and only the 4d cell of the fourth quartet contributes to larval structure, forming the longitudinal muscles, mesenchyme, and probably endoderm. Our results demonstrate a striking similarity between the cell lineages of polyclad and higher spiralian embryos, in which the four quadrants also bear the same relationships to the larval axes and give rise to comparable larval structures, including derivation of mesoderm from both ectodermal (2b) and endodermal precursors (4d)
gas2Is a Multifunctional Gene Involved in the Regulation of Apoptosis and Chondrogenesis in the Developing Mouse Limb

AbstractThe growth-arrest-specific 2 (gas2) gene was initially identified on account of its high level of expression in murine fibroblasts under growth arrest conditions, followed by downregulation upon reentry into the cell cycle (Schneideret al.,Cell 54, 787–793, 1988). In this study, the expression patterns of thegas2gene and the Gas2 peptide were established in the developing limbs of 11.5- to 14.5-day mouse embryos. It was found thatgas2was expressed in the interdigital tissues, the chondrogenic regions, and the myogenic regions. Low-density limb culture and Brdu incorporation assays revealed thatgas2might play an important role in regulating chondrocyte proliferation and differentiation. Moreover, it might play a similar role during limb myogenesis. In addition to chondrogenesis and myogeneis,gas2is involved in the execution of the apoptotic program in hindlimb interdigital tissues—by acting as a death substrate for caspase enzymes. TUNEL analysis demonstrated that the interdigital tissues underwent apoptosis between 13.5 and 15.5 days. Exactly at these time points, the C-terminal domain of the Gas2 peptide was cleaved as revealed by Western blot analysis. Moreover, pro-caspase-3 (an enzyme that can process Gas2) was cleaved into its active form in the interdigital tissues. The addition of zVAD-fmk, a caspase enzyme inhibitor, to 12.5-day-old hindlimbs maintained in organ culture revealed that the treatment inhibited interdigital cell death. This inhibition correlated with the absence of the Gas2 peptide and pro-caspase-3 cleavage. The data suggest that Gas2 might be involved in the execution of the apoptotic process
Characterization of Oocyte and Follicle Development in Growth Differentiation Factor-9-Deficient Mice

AbstractFemale mice null for the oocyte-specific gene product, growth differentiation factor-9 (GDF-9), a member of the transforming growth factor-β superfamily, exhibit primary infertility due to failed ovarian follicular development. The purpose of this study is to characterize oocyte and follicular differentiation as a function of animal age using cell culture and fluorescence, confocal, and electron microscopy. Analysis of follicles from GDF-9 homozygous mutant mice indicates that GDF-9-deficient oocytes grow more rapidly than control oocytes and that follicle growth ceases at the type 3b stage. Based on germinal vesicle (GV) chromatin patterns, fully grown oocytes isolated from GDF-9-deficient mice progress to advanced stages of differentiation equivalent to those found in antral follicles of control (heterozygous) mice.In vitromaturation of oocytes from homozygous mutant mice revealed that most oocytes are capable of resuming meiosis, with the ability to achieve meiotic completion reaching the highest levels in 6-week-old mice. Among the characteristic ultrastructural features of oocytes from homozygous mutant mice are perinuclear organelle aggregation, unusual peripheral Golgi complexes, and a failure to form cortical granules. Modified interconnections between granulosa cells and oocytes were also observed by ultrastructural (EM) and fluorescence microscopic analysis of follicles from GDF-9-deficient mice. These modifications included a decrease in the number of actin-based transzonal processes and modifications of microtubule-based projections that over time gave rise to invasion of the perivitelline space with eventual loss of oocyte viability. These cell–cell aberrations suggest a critical role for GDF-9 in the regulation of growth in preantral follicles through a mechanism involving bidirectional somatic cell–germ cell interactions
Induction of Optical Density Waves and Chemotactic Cell Movement inDictyostelium discoideumby Microinjection of cAMP Pulses

AbstractThe development of most multicellular organisms involves coordinated cell movement. The early aggregation of Dictyostelium cells has been shown to be mediated by chemotactic movement to propagating waves of cAMP. We have proposed that propagating waves of a chemoattractant, most likely cAMP, also control the movement of cells in mounds and slugs. We have now used periodic pressure injection of pulses of cAMP in the extracellular space of aggregation streams, mounds, and slugs to investigate whether these signals can be relayed and control cell movement, using quantitative digital time-lapse microscopy. Our major findings are (1) short (0.1 s) pulses of cAMP (107molecules) were able to elicit optical density (OD) waves in fields of aggregating amoebae. They propagate from the micropipet outward and interact with endogenous OD waves. (2) Periodic injection of cAMP pulses into aggregation streams blocked the pulses coming from the center and led to the rapid accumulation of cells downstream of the pipet around the pipet. (3) Injection of pulses of cAMP into mounds elicited OD waves, which propagated from the pipet outward and interacted with the endogenous waves, indicating that the same propagator carries them. (4) Periodic microinjection of cAMP in the prespore zone of slugs led to accumulation of anterior-like cells around the micropipet followed by tip formation. Furthermore, the cAMP signal could control the spacing of the endogenous sorting pattern. These results strongly support the hypothesis that the optical density waves observed during early development up to the mound stage represent propagating cAMP waves. They suggest furthermore that cAMP is the morphogen that controls cell movements in slugs
Heart Malformations in Transgenic Mice Exhibiting Dominant Negative Inhibition of Gap Junctional Communication in Neural Crest Cells

AbstractTransgenic mice were generated expressing an α1 connexin/β-galactosidase fusion protein previously shown to exert dominant negative effects on gap junctional communication. RNase protection analysis and assays for β-galactosidase enzymatic activity showed that the transgene RNA and protein are expressed in the embryo and adult tissues.In situhybridization analysis revealed that in the embryo, expression was predominantly restricted to neural crest cells and their progenitors in the dorsal neural tube, regions where the endogenous α1 connexingene is also expressed. Dye-coupling analysis indicated that gap junctional communication was inhibited in the cardiac neural crest cells. All of the transgenic lines were homozygote inviable, dying neonatally and exhibiting heart malformations involving the right ventricular outflow tract—the same region affected in the α1 connexinknockout mice. As in the knockout mice, the conotruncal heart malformations were accompanied by outflow tract obstruction. Histological analysis showed that this was associated with abnormalities in the differentiation of the conotruncal myocardium. These results suggest that the precise level of gap junctional communication in cardiac neural crest cells is of critical importance in right ventricular outflow tract morphogenesis. Consistent with this possibility is the fact that cardiac crest cells from the α1 connexinknockout mice also exhibited a greatly reduced level of gap junctional communication. These studies show the efficacy of a dominant negative approach for manipulating gap junctional communication in the mouse embryo and demonstrate that targeted expression of this fusion protein can be a powerful tool for examining the role of gap junctions in mammalian development
Age-Dependent Differences in the Effects of GDNF and NT-3 on the Development of Neurons and Glia from Neural Crest-Derived Precursors Immunoselected from the Fetal Rat Gut: Expression of GFRα-1in Vitroandin Vivo

AbstractNo enteric neurons or glia develop in the gut below the rostral foregut in mice lacking glial cell line-derived neurotrophic factor (GDNF) or Ret. We analyzed the nature and age dependence of the effects of GDNF and, for comparison, those of NT-3, on thein vitrodevelopment of the precursors of enteric neurons and glia. Positive and negative immunoselection with antibodies to p75NTRwere used to isolate crest-derived and crest-depleted populations of cells from the fetal rat bowel at E12, 14, and 16. Cells were typed immunocytochemically. GDNF stimulated the proliferation of nestin-expressing precursor cells isolated at E12, but not at E14–16. GDNF promoted the development of peripherin-expressing neurons (E12 ⪢ E14–16) and expression of TrkC. GDNF inhibited expression of S-100-expressing glia at E14–16. NT-3 did not affect cells isolated at E12, never stimulated precursors to proliferate, and promoted glial as well as neuronal development at E14–16. GFRα-1 was expressed both by crest- and non-crest-derived cells, although only crest-derived cells anchored GFRα-1 and GFRα-2 (GFRα-1 ⪢ GFRα-2). GDNF increased the number of neurons anchoring GFRα-1. GFRα-1 is immunocytochemically detectable in neurons of the E13 intestine and persists in adult neurons of both plexuses. We suggest that GDNF stimulates the proliferation of an early (E12) NT-3-insensitive precursor common to enteric neurons and glia; by E14, this common precursor is replaced by specified NT-3-responsive neuronal and glial progenitors. GDNF exerts a neurotrophic, but not a mitogenic, effect on the neuronal progenitor. The glial progenitor is not maintained by GDNF
Morphogenetic Mechanisms of Epithelial Tubulogenesis: MDCK Cell Polarity Is Transiently Rearranged without Loss of Cell–Cell Contact during Scatter Factor/Hepatocyte Growth Factor-Induced Tubulogenesis

AbstractMany organ systems are composed of networks of epithelial tubes. Recently, molecules that induce development of epithelial tubules and regulate sites of branching have been identified. However, little is known about the mechanisms regulating cell rearrangements that are necessary for tubule formation. In this study we have used a scatter factor/hepatocyte growth factor-induced model system of MDCK epithelial cell tubulogenesis to analyze the mechanisms of cell rearrangement during tubule development. We examined the dynamics of cell polarity and cell–cell junctions during tubule formation and present evidence for a multistep model of tubulogenesis in which cells rearrange without loss of cell–cell contacts and tubule lumens formde novo.A three-dimensional analysis of markers for apical and basolateral membrane subdomains shows that epithelial cell polarity is transiently lost and subsequently regained during tubulogenesis. Furthermore, components of cell–cell junctional complexes undergo profound rearrangements: E-cadherin is randomly distributed around the cell surface, desmoplakins I/II accumulate intracellularly, and the tight junction protein ZO-1 remains localized at sites of cell–cell contact. This suggests that differential regulation of cell–cell junctions is important for the formation of tubules. Therefore, during tubulogenesis, cell–cell adhesive contacts are differentially regulated while the polarity and specialization of plasma membrane subdomains reorganize, enabling cells to remain in contact as they rearrange into new structures
Delta–Notch Signaling in Odontogenesis: Correlation with Cytodifferentiation and Evidence for Feedback Regulation

AbstractRecent data suggest that dental cells utilize the evolutonarily conserved Notch-mediated intercellular signaling pathway to regulate their fates. Here we report on the expression and regulation of Delta1, a transmembrane ligand of the Notch receptors, during mouse odontogenesis.Delta1is weakly expressed in dental epithelium during tooth initiation and morphogenesis, but during cytodifferentiation, expression is upregulated in the epithelium-derived ameloblasts and the mesenchyme-derived odontoblasts. The expression pattern ofDelta1in ameloblasts and odontoblasts is complementary toNotch1, Notch2,andNotch3expression in adjacent epithelial and mesenchymal cells. Notch1 and Notch2 are upregulated in explants of dental mesenchyme adjacent to implanted cells expressingDelta1,suggesting that feedback regulation by Delta–Notch signaling ensures the spatial segregation of Notch receptors and ligands. TGFβ1 and BMPs induceDelta1expression in dental mesenchyme explants at the stage at whichDelta1is upregulatedin vivo,but not at earlier stages. In contrast to the Notch family receptors and their ligand Jagged1, expression ofDelta1in the tooth germ is not affected by epithelial–mesenchymal interactions, showing that the Notch receptors and their two ligands Jagged1 and Delta1 are subject to different regulations
In VitroStudies of Growth Cone Behavior Support a Role for Fasciculation Mediated by Cell Adhesion Molecules in Sensory Axon Guidance during Development

AbstractAxonal interactions, which are mediated by cell adhesion molecules (CAMs) as well as other types of membrane proteins, are important for sensory axon pathfinding in the developing chick hindlimb. We have previously shown that injection of antibodies that block the function of either G4/L1 or N-cadherin into the limb, starting when the first sensory axons reach the plexus, alters the segmental pattern of projections along cutaneous nerves. Specific removal of polysialic acid from NCAM using the enzyme endoneuraminidase N (Endo N) also resulted in significant changes in cutaneous projection patterns, while injection of antibodies against NCAM itself had no obvious effect (M. G. Honig and U. S. Rutishauser, 1996,Dev. Biol.175, 325–337). To help understand the cellular basis for these findings, we developed a tissue culture system in which the axons from dorsal root ganglion explants grow within defined laminin lanes and examined whether the same treatments increased or decreased a growth cone's tendency to be closely associated with neighboring axons. After 2 days in culture, images of the cultures were recorded, antibodies or Endo N was added, and images of the same fields were recaptured an hour later. To quantify the results, growth cones located in defined regions of the laminin lanes were classified, before and after the perturbation, as “free” (i.e., growing primarily on the laminin substratum), “fasciculated” (i.e., growing tightly along other neurites), or “intermediate” (i.e., growing both on the laminin substratum and in contact with other neurites). We found that anti-G4/L1 and anti-N-cadherin, but not anti-NCAM, caused an increase in defasciculated growth cones, whereas Endo N resulted in an increase in fasciculated growth cones. These changes in fasciculation are consistent with the changes in cutaneous projections seen in our previousin ovoperturbations. The results from these tissue culture experiments thus provide strong support for the idea that one mechanism by which CAMs affect sensory axon pathfindingin vivois by regulating the affinity of sensory growth cones for neighboring axons, which in turn can modulate the growth cone's ability to navigate through the surrounding environment
Regulation of Avian Cardiac Myogenesis by Activin/TGFβ and Bone Morphogenetic Proteins

AbstractPrevious studies have identified two signaling interactions regulating cardiac myogenesis in avians, a hypoblast-derived signal acting on epiblast and mediated by activin or a related molecule and an endoderm-derived signal acting on mesoderm and involving BMP-2. In this study, experiments were designed to investigate the temporal relationship between these signaling events and the potential role of other TGFβ superfamily members in regulating early steps of heart muscle development. We find that while activin or TGFβ can potently induce cardiac myogenesis in pregastrula epiblast, they show no capacity to convert noncardiogenic mesoderm toward a myocardial phenotype. Conversely, BMP-2 or BMP-4, in combination with FGF-4, can readily induce cardiac myocyte formation in posterior mesoderm, but shows no capacity to induce cardiac myogenesis in epiblast cells. Activin/TGFβ and BMP-2/BMP-4 therefore have distinct and reciprocal cardiac-inducing capacities that mimic the tissues in which they are expressed, the pregastrula hypoblast and anterior lateral endoderm, respectively. Experiments with noggin and follistatin provide additional evidence indicating that BMP signaling lies downstream of an activin/TGFβ signal in the cardiac myogenesis pathway. In contrast to the cardiogenic-inducing capacities of BMP-2/BMP-4 in mesoderm, however, we find that BMP-2 or BMP-4 inhibits cardiac myogenesis prior to stage 3, demonstrating multiple roles for BMPs in mesoderm induction. These and other published studies suggest a signaling cascade in which a hypoblast-derived activin/TGFβ signal is required prior to and during early stages of gastrulation, regulated both spatially and temporally by an interplay between BMPs and their antagonists. Later cardiogenic signals arising from endoderm, and perhaps transiently from ectoderm, and mediated in part by BMPs, act on emerging mesoderm within cardiogenic regions to activate or enhance expression of cardiogenic genes such as GATA and cNkx family members, leading to cardiac myocyte differentiation
ACaenorhabditis elegansHomologue ofhunchbackIs Required for Late Stages of Development but Not Early Embryonic Patterning

AbstractWe have cloned aCaenorhabditis eleganshomologue of theDrosophilagap genehunchback(hb) and have designated ithbl-1(hunchback-like).hbl-1encodes a predicted 982-amino-acid protein, containing two putative zinc-finger domains similar to those ofDrosophilaHunchback. The gene is transcribed embryonically, but unlike the maternally expressedDrosophila hb,its mRNA is not detected inC. elegansoocytes. Ahbl-1::gfpreporter is expressed primarily in ectodermal cells during embryonic and larval development. Double-stranded RNA-interference (RNAi) was used to indicatehbl-1loss-of-function phenotypes. Progeny ofhbl-1(RNAi)hermaphrodites exhibit a range of defects; the most severely affected progeny arrest as partially elongated embryos or as hatching, misshapen L1 larvae. Animals that survive to adulthood exhibit variably dumpy (Dpy), uncoordinated (Unc), and egg-laying defective (Egl) phenotypes, as well as defects in vulval morphology (Pvl). Abnormal organization of hypodermal cells and expression of a hypodermal marker inhbl-1(RNAi)animals suggests that most of the phenotypes observed could be due to improper specification of hypodermal cells. The pattern ofhbl-1expression is similar to that reported for the leech hunchback homologueLzf-2,suggesting that these proteins may have similar biological functions in diverse species with cellular embryos
Biphasic Subcellular Localization of the DAZL-Related Protein Boule inDrosophilaSpermatogenesis

AbstractTheDrosophila boulegene is expressed exclusively in the male germline and encodes an RNA binding protein closely related to the mammalian fertility factors encoded by theDAZ(Deleted in Azoospermia) andDAZL(DAZ-like) genes. Mutation ofbouleblocks both meiotic divisions. Differentiation nonetheless continues, resulting in tetraploid spermatids that fail to mature into sperm. We have found that Boule localizes premeiotically to a perinucleolar region and then translocates to the cytoplasm at the onset of meiosis. We show that deletion of the Y chromosomeks-1fertility locus eliminates Boule nuclear localization, although it does not perturb entry into meiosis. Based on these observations we propose that Boule acts in the cytoplasm to regulate the stability or translation of messenger RNA encoding an essential meiotic factor