pinecone/core-2020-05-10-deduplication huggingface dataset:
Origin of Interstitial Cells of Cajal in the Mouse Intestine

AbstractThe interstitial cells of Cajal (ICC) are found in a number of different locations in the gastrointestinal tract, where they form close associations with both muscle cells and nerve terminals. In this study we examined the embryological origin of ICC in the mouse intestine to determine whether they arise from the neural crest or from the intestinal wall. Segments of intestine were removed from embryonic mice either before or after the arrival of neural crest cells (the precursors of enteric neurons and glial cells) and transplanted under the renal capsule of host (adult) mice and allowed to develop for 18–41 days. In the mouse intestine, antibodies to c-kitprotein selectively label ICC at a variety of locations, and antibodies to the NK1 receptor (the receptor for substance P) labels ICC at the level of the deep muscular plexus in the small intestine and a subpopulation of enteric neurons in the large intestine. The presence of neurons in the explants was examined using antisera to neuron-specific enolase, substance P, and calretinin. In segments of small and large intestine explanted after the arrival of neural crest cells, immunoreactive neurons and c-kit- and NK1-immunoreactive ICC were present with a distribution similar to that seen in control tissue at a similar developmental age. In segments of large intestine explanted before the arrival of neural crest cells, neurons were not present; however, c-kit-immunoreactive ICC were present in these aneuronal explants, indicating that ICC do not arise from the neural crest. The source of ICC in mammals is therefore likely to be the mesenchyme of the gut
Fertilization Stimulates an Increase in Inositol Trisphosphate and Inositol Lipid Levels inXenopusEggs

AbstractPrevious experiments from our lab have suggested that the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) is required for sperm-induced egg activation inXenopus laevis.Here we measure the endogenous production of both Ins(1,4,5)P3and PIP2during the sperm-induced and ionomycin-induced calcium wave in the egg and find that both increase following fertilization. Ins(1,4,5)P3increases 3.2-fold from an unfertilized egg level of 0.13 pmole per egg (0.29 μM) to a peak of 0.42 pmole per egg (0.93 μM) as the calcium wave reaches the antipode in the fertilized egg. This continuous production of Ins(1,4,5)P3during the time that the Ca2+wave is propagating across the egg suggests the involvement of Ins(1,4,5)P3in wave propagation. This increase in Ins(1,4,5)P3is smaller in ionomycin-activated eggs than in sperm-activated eggs, suggesting that the sperm-induced production of Ins(1,4,5)P3involves a PIP2hydrolysis pathway that is not simply raising intracellular Ca2+. While one might expect PIP2levels to fall as a result of hydrolysis, we find that PIP2actually increases 2-fold. The total lipid fraction in unfertilized egg exhibits 0.8 pmole PIP2per egg and this increases to 1.5 pmole as the calcium wave reaches the antipode. The PIP2concentration peaks 2 min after the completion of the calcium wave at 1.8 pmole per egg. The amount of PIP2in the animal and vegetal hemispheres of the egg was also measured by cutting frozen eggs in half. The vegetal hemisphere contained twice the amount of PIP2as the animal hemisphere but it also contained twice the amount of lipid. Thus, there was an equivalent amount of PIP2normalized to lipid in each hemisphere. Isolated animal and vegetal hemisphere cortices exhibit similar PIP2concentrations, suggesting that the 2-fold higher total PIP2in the vegetal half is not due to a gradient of PIP2in the plasma membrane, but rather implies that cytoplasmic organelle membranes also contain PIP2
Elaboration of the Messenger Transport Organizer Pathway for Localization of RNA to the Vegetal Cortex ofXenopusOocytes

AbstractPrevious studies demonstrated that there were two pathways, the messenger transport organizer (METRO) or early and the Vg1 or late, which function during stages 1 to 3 of oogenesis for the localization of RNAs at the vegetal cortex ofXenopusoocytes. In the present study we analyzed the properties of the METRO pathway, which localizes Xlsirt, Xcat2, and Xwnt11 RNAs to a specific region of the vegetal cortex during stage 1 of oogenesis. A combination of methodologies involving both fixed material and living oocytes was used to analyze RNA localization. We show that in early diplotene pre-stage 1 oocytes (25–50 μm in diameter) both endogenous and injected exogenous METRO RNAs translocated to multiple mitochondrial aggregates (pre-mitochondrial clouds) that surround the germinal vesicle (GV). However, by early stage 1 (diplotene oocytes, 50–200 μm), all three of the RNAs discriminated between the different clouds and translocated exclusively within the METRO of a single mitochondrial cloud. Therefore, in stage 1 diplotene oocytes there is a unique mechanism causing a change in the intrinsic property of the mitochondrial clouds which designates one of them as the RNA transport vehicle. During translocation through the cytoplasm Xlsirt and Xcat2 RNAs were detected associated with cytoplasmic particles of different morphologies. Additionally, we also found that the translocation of RNAs through the early or METRO pathway, unlike that of the late pathway, occurred in the absence of intact microtubule and actin microfilament cytoskeletal elements. This supports a cytoskeletal-independent model for localization of RNAs through the METRO pathway
Identification of Novel Paired Homeodomain Protein Related toC. elegansunc-4 as a Potential Downstream Target of MASH1

AbstractA novel paired homeodomain protein, PHD1, that is most closely related toC. elegansunc-4 has been identified by a differential RT–PCR method.PHD1is expressed in a narrow layer adjacent to the ventricular zone of the dorsal spinal cord, immediately following expression of MASH1 but preceding overt neuronal differentiation. Some cells coexpressing MASH1 andPHD1can be seen, suggesting that these two genes are sequentially activated within the same lineage. In the olfactory sensory epithelium,PHD1expression not only follows but is dependent upon MASH1 function, suggesting that PHD1 acts downstream of MASH1. A sequential action of bHLH and paired homeodomain proteins is apparent in other neurogenic lineages and may be a general feature of both vertebrate and invertebate neurogenesis
The Inhibition of Differentiation Caused by TGFβ in Fetal Myoblasts Is Dependent upon Selective Expression of PKCθ: A Possible Molecular Basis for Myoblast Diversification during Limb Histogenesis

AbstractEmbryonic and fetal skeletal myoblasts are responsible for the formation of primary and secondary fibers in mammals, but the mechanism which diversifies their fate is unknown.In vitro,embryonic myoblasts are resistant to the differentiation inhibitory effects of transforming growth factor β and phorbol esters. Thus, differential expression of specific molecules involved in the transduction of extracellular signals may contribute to the different phenotypes. We report here that protein kinase C θ, but none of the other known protein kinase C isoforms, is selectively expressed in fetal and postnatal muscle cells (at both the myoblast and myotube stage)in vitroandin vivo.By contrast, embryonic myoblasts and myotubes do not express protein kinase C θin vitroorin vivo.This difference is causally related to a differential response to transforming growth factor β, since overexpression of protein kinase C θ, but not of protein kinase C α or ζ, in embryonic myoblasts makes these cells sensitive to transforming growth factor β. These data demonstrate for the first time that a protein kinase C isoform is a key component of the signal transduction cascade which follows exposure of myoblasts to transforming growth factor β. They also suggest a specific role for protein kinase C θ in determining the fate of different myoblasts during muscle histogenesis
A Cluster of Noninvoluting Endocytic Cells at the Margin of the Zebrafish Blastoderm Marks the Site of Embryonic Shield Formation

AbstractIn zebrafish embryos, the nascent embryonic shield first appears as a thickening in the germ ring of the mid-epiboly blastoderm. This site defines the dorsal side of the developing embryo. In this paper, we report that the site of embryonic axis formation is marked earlier at the late-blastula stage by the appearance of a cluster of cells with unique endocytic activities. This cluster of cells is composed of enveloping layer epithelial cells and one to two layers of underlying deep cells. Unlike other marginal blastomeres, cells in this cluster do not participate in involution as the blastoderm undergoes epiboly. These noninvoluting endocytic marginal (NEM) cells can be selectively labeled by applying membrane impermeant fluorescent probes to pre-epiboly and mid-epiboly embryos. During embryonic shield formation, deep cells in the NEM cell cluster rearrange and are displaced forward to the leading edge of the blastoderm. As deep NEM cells move into this location, they become a group of cells known as “forerunner cells.” Between 60%- and 80%-epiboly, the forerunner cells coalesce into a coherent cell cluster that forms a wedge-shaped cap at the leading edge of the blastoderm. During embryonic axis formation, deep cells migrate and converge toward the embryonic midline, which is defined by the center of the forerunner cell cluster. At approximately 90% epiboly, the forerunner cell cluster becomes overlapped by the constricting germ ring. At tailbud stage, forerunner cells form the dorsal roof of Kupffer's vesicle, which is located ventral to the nascent chordoneural hinge. On the basis of previous grafting studies and known dorsal gene expression patterns, we discuss possible roles that the NEM/forerunner cell cluster may play in teleost axis formation
Alteration of Cell Cycle Timing and Induction of Surface Instability in Starfish Blastomeres Microinjected with Antibodies to Spectrin

AbstractSpectrin has been implicated in a variety of different processes during late embryogenesis, after transcription of the zygotic genome has been activated. However, relatively little is known about the role of maternally derived spectrin during the early cleavage divisions that give rise to a multicellular embryo. To investigate the role of spectrin in early development, we have microinjected anti-spectrin antibodies intoPatiria miniatastarfish embryos to inhibit the activity of the maternal pool of spectrin. Microinjection of affinity-purified anti-spectrin antibody, or low to moderate doses of Fabfragments, into one blastomere of a two-cell-stage embryo caused a dose-dependent, progressive increase in the length of the cell cycle compared to the uninjected control blastomere. The progeny of injected blastomeres were unable to participate in the formation of a blastula epithelium, instead forming a loose aggregate of cells that eventually stopped dividing. When division stopped, the cells formed surface protrusions and became motile. At high doses of either whole antibody or Fabfragments, cells initiated, but failed to complete, cytokinesis. Blastomeres injected with high doses of Fabfragments also failed to reform nuclei and underwent variable periods of cell cycle arrest up to 12 hr. Injected embryos stained with BODIPY-phallacidin exhibited extensive disruption of the cortical actin cytoskeleton. These results support previous studies implicating spectrin in stabilizing the cell surface and maintaining the organization of the cortical cytoskeleton. They further suggest that spectrin is not required for the initiation or contraction of the cleavage furrow, but functions in the completion of cytokinesis. Most surprisingly, however, the results demonstrate that inhibition of spectrin function alters cell cycle timing, suggesting that disruption of the actin cytoskeleton inhibits progression through the cell cycle
Activation of thereaperGene during Ectopic Cell Killing inDrosophila

AbstractThe product of thereaper(rpr) gene is required for programmed cell death inDrosophila.We examinedrprexpression during ectopic cell deaths caused by ionizing radiation or aberrant development. In both instances, dramatic induction ofrprexpression was observed. A genomic fragment upstream ofrprconfers this regulatory behavior upon alacZreporter transgene. In a model cell culture system, conditional expression of REAPER is sufficient to induce massive apoptosis that can be prevented by the anti-apoptotic protein p35. Overall, these results suggest that diverse signals converge at, or upstream of,rpr-associated transcriptional regulatory elements that can function to initiate a common apoptotic pathway involving ICE-like protease activity
Restricted Expression of a Novel Murineatonal-Related bHLH Protein in Undifferentiated Neural Precursors

AbstractTissue-specific bHLH proteins play important roles in the specification and differentiation of neural cell lineages in invertebrate and vertebrate organisms. Two groups of bHLH proteins, atonal and achaete-scute, have proneural activities inDrosophila,and the mouse achaete-scute homolog MASH1 is required for the differentiation of several neural lineages. In a screen for proteins interacting with MASH1, we have isolated a novel bHLH protein related to atonal, named MATH4A, which is broadly expressed in neural precursor cells in the mouse embryonic CNS and PNS. Interaction assays in yeast andin vitrodemonstrate that MATH4A interacts efficiently with both MASH1 and the ubiquitous bHLH protein E12. MATH4A-E12 heterodimers, but not MATH4A-MASH1, bind to a consensus E-box sequence.Math4Aexpression is restricted to undifferentiated neural precursors and is complementary to that ofMash1in most regions of the nervous system. In particular,Math4Ais transcribed at high levels in the cerebral cortex, dorsal thalamus, and epibranchial placodes, which present little or noMash1expression. However, expression of the two genes shows limited overlap in certain CNS regions (retina, preoptic area of the hypothalamus, midbrain, hindbrain). Its structure and expression pattern suggest that MATH4A may regulate an early step of neural development, either as a partner of ubiquitous bHLH proteins or associated with other neural-specific bHLH proteins
Abrogation of Transforming Growth Factor-β Type II Receptor Stimulates Embryonic Mouse Lung Branching Morphogenesis in Culture

AbstractTGF-β1 is a known inhibitor of branching morphogenesis when added exogenously to mouse embryonic lungs in culture. However, the issue of whether endogenous TGF-β signaling has a function in the process of lung organogenesis is not completely resolved. We utilized immunoperturbation and antisense oligodeoxynucleotide inhibitory strategies to abrogate TGF-β type II receptor function in embryonic mouse lungs undergoing branching morphogenesis in serumless explant culture. Antisera directed against a TGF-β type II receptor N-terminal peptide that perturbs TGF-β ligand–receptor binding increased branching by 70%. Similarly, antisense TGF-β type II receptor oligodeoxynucleotides (40 μM) resulted in a 58% increase in branching, compared to scrambled and mismatched sequence controls, while TGF-β type II receptor mRNA and its protein expression levels were suppressed by 95 and 84%, respectively. Addition of exogenous TGF-β1 did not overcome the stimulatory effects either of TGF-β type II receptor immunoperturbation or of antisense oligodeoxynucleotide treatment on lung branching morphogenesis. Usingin situhybridization and immunohistochemistry, both TGF-β type II receptor mRNA and protein were localized to the epithelium lining the developing airways, and to the surrounding mesenchyme, indicating that TGF-β type II receptor is an important regulator of epithelial–mesenchymal interaction. Exogenous TGF-β1 decreased cyclin A mRNA levels in control embryonic lung explants, while TGF-β type II receptor antisense oligodeoxynucleotides prevented the downregulation of cyclin A mRNA expression by exogenous TGF-β1. In addition, PCNA immunostaining of the primitive bronchial epithelium was increased in the presence of TGF-β type II receptor antisense oligodeoxynucleotides either alone or together with exogenous TGF-β1, whereas TGF-β1 alone decreased PCNA staining. Thus, abrogation of TGF-β type II receptor expression prevented TGF-β1-induced epithelial cell G1arrest. These results demonstrate, for the first time, that abrogation of the TGF-β type II receptor stimulates embryonic lung organogenesis in culture and reverses the negative influence of endogenous TGF-β signaling upon epithelial cell cycle progression
Developmental Profiles and Ecdysteroid Regulation of the mRNAs for Two Ecdysone Receptor Isoforms in the Epidermis and Wings of the Tobacco Hornworm,Manduca sexta

AbstractEcdysteroids acting through multiple isoforms of the ecdysone receptor (EcR) initiate molting and metamorphosis of insects. Two isoforms of EcR, A (this paper) and B1 (Fujiwaraet al., Insect Biochem. Mol. Biol.25, 845–856, 1995), were isolated from the tobacco hornworm,Manduca sexta,and shown to be similar to the correspondingDrosophilaEcR isoforms. The developmental profiles of both EcR-A and EcR-B1 (determined by both analysis of isoform-specific mRNAs and use of monoclonal antibodies that detect either EcR-B1 or all forms), however, were different inManducaepidermis, which produces sequentially the larval, the pupal, and the adult cuticles. EcR-B1 predominated through the larval, pupal, and early developing adult stages with an upregulation early in each molt. By contrast, EcR-A was present only at the onset of new cuticle synthesis during the larval molt, but in the pupal and adult molts was upregulated slightly later than EcR-B1 during the commitment period and was present during the predifferentiative phase. Both isoforms appeared in the larval wing discs after pupal commitment and persisted through pupal differentiation. The mRNAs for both isoforms were directly induced in larval epidermisin vitroby 20-hydroxyecdysone, but EcR-B1 mRNA accumulated more rapidly, peaking at 3 hr. In the presence of a protein synthesis inhibitor, the accumulation of EcR-B1 mRNA was slower and its subsequent decline was prevented, but the accumulation of EcR-A mRNA was unaffected. Thus, in this polymorphic epidermis both isoforms appear in every molt, with EcR-B1 present during the commitment and predifferentiative phases and then at the onset of cuticle synthesis EcR-A prevails. Additionally, EcR-A is apparently associated with the switching and predifferentiative events necessary for a new synthetic program
Sonic hedgehog Differentially Regulates Expression ofGLIandGLI3during Limb Development

AbstractSonic hedgehog is a secreted factor regulating patterning of the anterior–posterior axis in the developing limb. The signaling pathway mediating the transduction of the signal is still poorly understood. InDrosophilaseveral genes are known to act downstream ofhedgehog,the fly homolog ofSonic hedgehog.An important gene epistatic tohedgehogiscubitus interruptus,which encodes the fly homolog of a family of vertebrate putative transcription factors, theGLIgenes. We have isolated two members of theGLIfamily from chick, calledGLIandGLI3.Their expression patterns in a variety of tissues during embryogenesis suggest that these genes may be targets of the Sonic hedgehog signal. We demonstrate that the twoGLIgenes are differentially regulated by Sonic hedgehog during limb development. Sonic hedgehog up-regulatesGLItranscription, while down-regulatingGLI3expression in the mesenchymal cells of the developing limb bud. Finally, we demonstrate that an activated form of GLI can induce expression ofPatched,a known target of Sonic hedgehog, thus implicating GLI as a key transcription factor in the vertebrate hedgehog signaling pathway. In conjunction with evidence from a mouseGli3mutant, our data suggest that GLI and GLI3 may have taken two different functions of theirDrosophilahomologcubitus interruptus
Regulation of Protein Tyrosine Phosphorylation in Human Sperm by a Calcium/Calmodulin-Dependent Mechanism: Identification of A Kinase Anchor Proteins as Major Substrates for Tyrosine Phosphorylation

AbstractSignal transduction pathways regulate various aspects of mammalian sperm function. When human sperm were incubated in a medium supporting capacitation, proteins became tyrosine-phosphorylated in a time-dependent manner. This phosphorylation was inhibited by genistein, a protein tyrosine kinase inhibitor. Phosphorylation was also reduced when sperm were incubated either in the presence of increasing concentrations of extracellular Ca2+or in a medium containing the Ca2+ionophore A23187. This Ca2+-induced dephosphorylation was calmodulin-dependent, suggesting that calcineurin was involved. In this regard, the calcineurin inhibitor deltamethrin inhibited the Ca2+ionophore-induced dephosphorylation. A limited number ofMr80,000–105,000 polypeptides were the most prominent phosphotyrosine-containing proteins present in human sperm. Unlike mouse sperm, which contains a tyrosine-phosphorylated isoform of hexokinase, a phosphotyrosine-containing hexokinase in human sperm was not detected. Most of the tyrosine-phosphorylated proteins were Triton X-100-insoluble and were localized to the principal piece of the flagellum, the region where the cytoskeletal fibrous sheath is found. Prominent phosphotyrosine-containing proteins ofMr82,000 and 97,000 were identified as the human homologues of mouse sperm AKAP82, the major fibrous sheath protein, and pro-AKAP82, its precursor polypeptide, respectively. These proteins are A Kinase Anchor Proteins, polypeptides that sequester protein kinase A to subcellular locations. Taken together, these results suggest that protein tyrosine phosphorylation may be part of a signal transduction cascade(s) regulating events pertaining to capacitation and/or motility in mammalian sperm and that an interrelationship between tyrosine kinase and cAMP signaling pathways exists in these cells
Perturbation of the DevelopingXenopusRetinotectal Projection Following Injections of Antibodies against β1Integrin Receptors and N-cadherin

AbstractWe have examined the function of β1integrin receptors and N-cadherin in the development of theXenopusretinotectal projection.In vivoperturbation experiments were performed by injecting antibodies directed against β1integrin receptors and N-cadherin into the embryonic optic pathway. The antibodies were present during the initial development of the retinal projection, when the axons of the ganglion cells are migrating through the optic tract and terminating within the optic tectum. When injected individually, the antibodies were insufficient to cause obvious pathfinding errors. However, when injected together, the antibodies caused specific abnormalities in the development of the retinotectal projection. Pathfinding errors most commonly observed included ectopically projecting axons within the optic tract region, meandering and splaying of axons in the optic tectum, and the induction of prominent ipsilateral projections. IgGs and Fab′ fragments of the antibodies produced pathfinding errors; these defects were not observed in animals injected with control antibodies. Thesein vivoresults show that β1integrin receptors and N-cadherin have important roles during the development of the visual projection and provide evidence that a balance between cell–cell and cell–matrix adhesion may be critical for the normal development of the vertebrate visual system