pinecone/core-2020-05-10-deduplication huggingface dataset:
The Ames Dwarf Gene Is Required for Pit-1 Gene Activation

AbstractThe tissue-specific POU domain transcription factor Pit-1 is required for phenotypic specification of three cell types in the anterior pituitary gland of mammals: somatotropes, thyrotropes, and lactotropes. Mutations in thePit-1gene, as first described in Snell and Jackson dwarf (dw) mice, led to pituitary hypoplasia due to lack of these three cell types, as well as hypothyroidism and dwarfism because of deficiency of thyroid-stimulating hormone and growth hormone, respectively. The Ames dwarf (df) exhibits a phenotype identical to that of Pit-1-mutated mice but the defective gene has been mapped to a locus on mouse chromosome 11, distinct from thePit-1gene located on mouse chromosome 6. Our studies indicate that initial activation of thePit-1gene is deficient in the Ames dwarf. This suggests that thedfgene is required for activation of thePit-1gene and provides evidence for a hierarchy of tissue-specific factors required for cellular commitment in the anterior pituitary gland
Heterochrony and the Phylotypic Period

AbstractThere has been a resurgence of interest in comparative embryology. It is now important to be able to compare gene expression in different species at similar developmental stages. One phenomenon which may make it difficult to compare embryos in this way isheterochrony—a change in developmental timing during evolution. It is not clear whether heterochrony can affect the intermediate stages of embryonic development, when many important genes involved in pattern formation are expressed. A prevalent view is that these so-called phylotypic stages are resistant to evolutionary change because they are when the body plan is laid down. Haeckel's famous drawings, which show different vertebrates developing from virtually identical somite-stage embryos, are still used to support this idea. I have reexamined the morphological data relating to developmental timing in somite-stage embryos. The data reveal striking patterns of heterochrony during vertebrate evolution. These shifts in developmental timing have strongly affected the phylotypic stage, which is therefore poorly conserved and is more appropriately described as the phylotypic period. This is contrary to the impression created by Haeckel's drawings, which I show to be inaccurate and misleading. The study of gene expression in embryos which show heterochrony could give important insights into evolutionary and developmental mechanisms
A Developmental Role for the Heterotrimeric G Protein Goαin a Migratory Population of Embryonic Neurons

AbstractThe heterotrimeric G proteins are an extended family of guanyl nucleotide-binding proteins that serve essential functions in the mature nervous system but whose contributions to neuronal development remain poorly understood. We have investigated the potential role of one specific G protein, Goα, in the control of neuronal migration. During embryogenesis of the moth,Manduca sexta,an identified population of undifferentiated neurons (the EP cells) migrate along sets of visceral muscle bands to form part of the enteric nervous system. Previously, immunohistochemical studies indicated the presence of Goα-related proteins in the EP cells during migration. We have now verified this result, using probes derived from the Goαgene inManduca.A clone containing the full-length coding domain for Goαwas sequenced from aManducacDNA library; digoxigenin-labeled probes were then made from this clone and used to examine the developmental expression of the Goαgene during embryogenesis. Goα-specific transcripts could first be detected in the EP cells several hours before the onset of their migration. The level of Goαexpression in all of the EP cells continued to increase during migration, but subsequently was down-regulated in a subset of the postmigratory neurons at the time of their terminal differentiation. This pattern of regulated expression is consistent with the distribution of Goα-related protein in the EP cells. We also used a semi-intact culture preparation of staged embryos to investigate the effects of G protein-specific toxins on the migratory process. Intracellular injections of the wasp toxin mastoparan, a specific activator of Goα- and Giα-related proteins, inhibited the migration of individual EP cells. Injections of pertussis toxin (an inhibitor of Goαand Giα) or cholera toxin (a selective activator of Gsα) had no effect on migration, although pertussis toxin treatments did cause a measurable increase in the subsequent outgrowth of axonal processes. However, co-injection of mastoparan with pertussis toxin blocked the inhibitory effects of mastoparan alone. These results suggest that Goα-coupled signaling events within the EP cells may down-regulate their migratory behavior, possibly in response to inhibitory cues that normally guide migration in the developing embryo
Propagated and Nonpropagated Calcium Transients during Egg Activation in the Annelid,Chaetopterus

AbstractTransient waves of Ca2+release cross-fertilizing deuterostome eggs from the point of sperm entry to its antipode and provide much of the activating stimulus for the egg. Based on several indirect lines of experimental evidence, it was proposed that protostome eggs are activated by a prolonged uptake of Ca2+from the medium due to sperm-induced membrane depolarization and that this uptake then starts an activation wave similar to those in deuterostomes, except that it moves inward from the whole surface rather than through the egg from pole to pole. To test these hypotheses, we microinjected oocytes of the polychaete annelid,Chaetopterus pergamentaceus,with semisynthetic recombinant aequorins and measured light emission in response to both fertilization and artificial activation by excess K+. Both fertilization and K+-activation induced multiple, brief Ca2+transients in the eggs. The first transient did not propagate, but it was followed by a series of globally propagated Ca2+waves interspersed with additional nonpropagated pulses. The waves traversed the egg at about 30 μm/sec. Sequential propagated waves and nonpropagated pulses generally originated at different regions on the egg surface, except the last few, which originated in the same “pacemaker” region. These new data are consistent with the hypothesis that the activation of protostome eggs is initiated by Ca2+waves. However, the fact that these waves propagated from pole to pole like those in deuterostome eggs refutes the notion that Ca2+waves in activating protostome eggs move inward from the whole surface
Four-Dimensional Microscopic Analysis of the Filopodial Behavior of Primary Mesenchyme Cells during Gastrulation in the Sea Urchin Embryo

AbstractDuring gastrulation of the sea urchin embryo, primary mesenchyme cells (PMCs) migrate from the vegetal pole to a site below the equator of the embryo where they form a ring-like structure and begin producing the larval skeleton. As these cells migrate, they extend and retract filopodia which appear to interact with the basal lamina and underlying ectoderm. To better characterize this behaviorin vivo,we studied PMC migration using differential interference contrast (DIC) microscopy in combination with four-dimensional imaging (x,y,zspace and time). We were able to determine the persistence and direction of extension of each filopodium and were also able to observe the dynamic behavior of each using colorized movie loops. This analysis showed that: (1) Most filopodia are quite transient, usually persisting for less than 0.5–6.0 min, during which time they continuously survey their surroundings; (2) PMCs extend an average of 121 filopodia/hr during migration; (3) the initial direction of extension of filopodia from the cell body is random, with just as many filopodia projecting toward as away from the direction of migration; (4) as a consequence of (2) and (3) above, each PMC explores the area surrounding its cell body approximately once every 5 min; (5) PMCs nearer to the target site migrate faster than those located farther away. To further investigate filopodial distribution, confocal microscopy was used to collectzseries of PMCs transplanted to different locations in the embryo and fixed during migration. We found that more filopodia tended to be distributed toward the target site as cells approached the ring, suggesting that filopodial distribution may reflect regional differences in directional cues
Translational Control in Spermatogenesis

AbstractTranslational silencing phenomena during spermatogenesis in the two model systemsDrosophilaand mouse are reviewed.Cis-acting sequences were identified in both species that are necessary for translational repression. While inDrosophilathese elements so far have only been found in the 5′ untranslated region (5′ UTR), in mammals such regions were identified both in the 5′ as well as in the 3′ UTR. In all cases, RNA-binding proteins interact with these regions, yet their specific role in the observed negative regulation of translation has to be established
Developmental Role of Transcription Factor Isoforms Generated by Alternative Splicing

AbstractThe production of transcription factor isoforms by developmentally regulated alternative splicing of pre-mRNAs is a widespread phenomenon. Frequently, differences in biochemical function among the isoforms can be predicted from sequence analysis, and in many instances such differences have been demonstratedin vitroor in cultured cells. A great variety of strategies for functional diversification can be classified into three main types: modulation of DNA binding specificity or affinity, production of activators and antagonists from the same gene, and modulation of dimerization properties. Despite obvious implications in many cases, the actual developmental consequences are understood only in a few instances. The roles inferred from these examples are diverse, ranging from developmental switches that have profound effects on pathways of differentiation to mechanisms that may optimize or fine-tune transcription factor function in different contexts
A Novel TGF-β-like Gene,fugacin,Specifically Expressed in the Spemann Organizer ofXenopus

AbstractUsing a differential screening strategy, we have cloned a novelXenopusgene,fugacin,related to the transforming growth factor β superfamily. Transcripts were detected primarily in the dorsal marginal zone of late blastula. Thereafter, they became highly localized to the blastopore lip of early gastrula and were not observed at later stages. This gene, which is most homologous to the mouse genenodal,displays a new pattern of cysteine residues. These findings highlight the potential role of these growth factors during early vertebrate development
trkC-Mediated NT-3 Signaling Is Required for the Early Development of a Subpopulation of Neurogenic Neural Crest Cells

AbstractSoon after they segregate from the neural tube, trunk neural crest cells disperse on two spatially and temporally distinct pathways. Only crest cells that migrate early and ventromedially give rise to neurons of the peripheral nervous system. It is also known that neural crest cell-derived populations require appropriate environmental cues early in development in order to generate neurons, and for the subsequent survival of differentiated neurons. We examined whether neurotrophin-3 (NT-3), a survival factor for subsets of peripheral neurons, is also involved in the regulation of neurogenesis by neural crest cells. First, we found that premigratory and migrating neural crest cells on the medial migration pathway of Embryonic Day 2.5 (E 2.5) embryos express mRNAs encoding multiple isoforms of the NT-3 receptor,trkC, as do cells in the neural tube and epithelial dermamyotome. On E4, a subpopulation of neurons in nascent sensory ganglia expresstrkC message. Second, we demonstrate thattrkC mRNA is only expressed in neural crest cell populations that possess neurogenic potential. Third, we show that the presence of NT-3, during the initial development of cultured neural crest cells, is required for neurogenesis by a subpopulation of neurogenic neural crest-derived cells. These results suggest that a subpopulation of neurogenic neural crest cells expresses functionaltrkC receptors and requires the timely availability of NT-3 for their development before reaching their final embryonic locations. We suggest that developmental heterogeneity exists in the identity and requirements of neural crest cell subsets that harbor neurogenic potential. We also suggest that the “paradoxical” expression oftrkC receptors by the somitic dermamyotome may, in fact, play a role in the exclusive development of crest-derived neurogenic precursors on the medial pathway by limiting the availability of NT-3 on the lateral pathway
Analysis of Neural-Responsive Myogenin Upstream Sequences by Myoblast Implantation

AbstractThe expression of myogenin is suppressed during innervation and has been implicated in determining properties of skeletal muscle which are regulated by electrical activity. We previously reported that transcription driven by 3700 bp of the mouse myogenin upstream sequence (MYG3700) is activated by denervation in transgenic mice (Nucleic Acids Res.21, 5684–5693, 1993). To extend our investigation of the activity dependence of the myogenin promoter, we have utilized myoblast implantation as a novel approach toin vivoreporter analysis. Myoblasts for hindlimb injections were generated by stable transfection of chloramphenicol acetyltransferase (CAT) reporters into a β-galactosidase-expressing line of C2 cells.In vitrocharacterization of stable myoblast clones carrying myogenin–CAT deletion constructs revealed that while the proximal myogenin 5′-flanking sequence confers myotube specificity, high-level expression requires a region upstream (−335 to −1102) which depends on chromosomal integration for its function. For analysis by implantation, incorporation of injected myoblasts into existing myofibers was confirmed by histochemical staining. Using clonal myoblasts harboring nicotinic receptor α-subunit (α800) and myosin light chain reporters as positive and negative controls, respectively, for denervation responsiveness, we determined that the nuclei of injected myoblasts are susceptible to regulatory signals imposed by nerve-induced electrical activity of the myofiber into which they incorporate. Inin vivoanalysis of myogenin upstream sequence by implantation, CAT activities of MYG3700 and MYG1565 reporters in injected limbs increased up to fourfold within 4 days after denervation, whereas the activities of MYG1102 and MYG335 were unchanged. By 10 days after denervation, all myogenin reporters displayed denervation responsiveness. These implantation data suggest an early phase of denervation activation, one that is mediated by control elements residing within −1102 to −1565 of the myogenin upstream sequence. Thus, the combined analyses of stable reporter myoblast linesin vitroandin vivoby implantation provide an efficient means of evaluating regulatory regions for high-level expression and neural modulation of muscle gene transcription
Expressed Ryanodine Receptor Can Substitute for the Inositol 1,4,5-Trisphosphate Receptor inXenopus laevisOocytes during Progesterone-Induced Maturation

AbstractTwo structurally related forms of intracellular calcium release channels that can mediate the release of intracellular calcium have been identified: the ryanodine receptors (RyR) and the inositol 1,4,5-trisphosphate receptors (IP3R). Each channel responds to distinct pathways for activation. The IP3R is activated by IP3 and the RyR is thought to be activated by calcium or by another second messenger cADP ribose. It has been proposed that each type of channel subserves a specialized pool of intracellular calcium, and it is not understood why some cell types require more than one form of intracellular calcium release channel. The present study was designed to examine whether the RyR can substitute for the IP3R during oocyte maturation. IP3R expression was inhibited inXenopus laevisoocytes using antisense oligonucleotides. These oocytes, with reduced levels of IP3R, demonstrated a marked delay in the time course of progesterone-induced maturation. The cloned skeletal muscle RyR1 was then expressed inX. laevisoocytes that were deficient in IP3R. Functional studies showed that the properties of the cloned RyR1, expressed in oocytes, were comparable to those of the native RyR1.X. laevisoocytes deficient in IP3R, but expressing RyR1, were able to undergo progesterone-induced maturation with a time course comparable to that seen in wild-type oocytes when caffeine was used to activate RyR and induce intracellular calcium release. These studies show that RyR1 can substitute for the IP3R as the intracellular calcium release channel required forXenopusoocyte maturation and that intracellular calcium release is important for controlling the rate of progesterone-induced maturation
An ECM-Bound, PDGF-like Growth Factor and a TGF-α-like Growth Factor Are Required for Gastrulation and Spiculogenesis in theLytechinusEmbryo

AbstractGrowth factors and the extracellular matrix have been shown to fulfill vital developmental roles in many embryonic systems. Our hypothesis is that a developmental role played by the extracellular matrix in sea urchins may be the binding of a PDGF-like growth factor to promote signaling activity. We report here that anti-human PDGF-B antibodies and anti-human TGF-α antibodies immunoprecipitated specific proteins isolated fromLytechinusembryos. Addition of these antibodies toLytechinusembryos inhibited gastrulation and spiculogenesis. The embryos are sensitive to the antibodies from the four-cell through the hatching blastula stages, which suggests that the TGF-α-like and PDGF-like ligands are required for the early differentiation of the gut and spicules. We present evidence that the PDGF-like growth factor depends on the extracellular matrix for signaling activity. Synthetic peptides representing the heparan sulfate proteoglycan binding sequence on human PDGF-B were added toLytechinusembryo cultures to compete for binding sites with the endogenous PDGF-like growth factor. The experimental peptide inhibited gastrulation and caused radially arranged multiple spicules to form. Development was unaffected by a control peptide. These studies support our hypothesis and suggest that TGF-α-like and PDGF-like growth factors induce signaling events required for sea urchin gastrulation and spiculogenesis and suggest that an extracellular matrix-associated PDGF-like growth factor is involved in differentiation along the oral–aboral axis
Imaging Protein Kinase C Activation in Living Sea Urchin Eggs after Fertilization

AbstractThe fluorescent dye NBD–phorbol acetate was used to visualize the activation of protein kinase C (PKC) in livingLytechinus pictuseggs during fertilization. This dye interacts directly with PKC as determined using a competitive binding assay. Quantitative image analysis of sequential images from laser-scanning confocal microscopy showed a significant reorganization of the signal in the vicinity of the cortical granules and the plasma membrane that began immediately following fertilization and persisted up to 1 hr (P< 0.0001). At the concentrations employed, the NBD–phorbol dye was not capable of inducing a significant translocation of the fluorescent signal to the membrane, nor did it appear to interfere with the cell cycle. It therefore seems likely that the presentin vivoresults reflect the previously reportedin vitroactivation of protein kinase C immediately subsequent to fertilization. Such an interpretation is parsimonious with the results of parallel subcellular fractionation experiments using an N-terminal polyclonal antibody to sea urchin PKC which showed a significant (P< 0.037) translocation of the enzyme from the cytosolic fraction to the membrane fraction 40 min subsequent to fertilization. This study supports and extends previousin vitrodata suggesting that PKC activation subsequent to fertilization occurs at or near the egg plasma membrane, perhaps in association with arachadonic acid-rich cortical granules
The Epidermis Is a Source of Directional Information for the Migrating Pronephric Duct inAmbystoma mexicanumEmbryos

AbstractIn the urodeleAmbystoma mexicanum,the pronephric duct (PND) is formed from a coherent group of cells that migrate from the pronephros to the cloaca along a pathway immediately ventral to the developing somites. The guidance cues used by the migrating PND primordium to find the cloaca are a local property of the migration substratum, are temporally regulated, and are both polarized and oriented. Since the pronephric duct migrates between two tissues—the underlying lateral mesoderm and the overlying epidermis—we performed a study to identify the tissue(s) in which PND guidance cues originate. Through direct manipulation of the epidermis overlying the duct pathway, we show that the migrating PND reads epidermally derived cues (1) along the anterior–posterior axis that direct migration from anterior to posterior and (2) along the dorsal–ventral axis that constrain migration to the duct pathway. Heterochronic grafting experiments reveal that the ability to direct PND migration is a stable property of flank epidermis throughout the period of PND migration. Epidermal cues are, therefore, not responsible for the observed temporal restrictions on PND migration. Thus, the region of the embryo within which the advancing PND tip can migrate actually represents an area where two distinct but required sets of PND migration cues overlap. The epidermis overlying the duct pathway provides directional information; temporal restriction of duct migration is hypothesized to be a property of the flank mesoderm
Changes in Responsiveness to Extracellular ATP in Chick Skeletal Muscle during Development and upon Denervation

AbstractSkeletal muscles in developing chick embryos were tested for responsiveness to adenosine 5′-triphosphate (ATP), a substance known to depolarize chick skeletal muscle in culture. The sensitivity to extracellular ATP was tested at various stages of development in five different muscles; pectoralis superficia, anterior latissimus dorsi, posterior latissimus dorsi, sartorious, and gastrocnemius. At the earliest time that muscles were tested (Embryonic Day 6, stage 30 of Hamburger and Hamilton, 1951) application of ATP (50–100 μM) elicited vigorous contractions in all five muscles, but within a few days (Embryonic Day 17, stage 43) none of the muscles contracted in response to ATP. Sensitivity declined at approximately the same time in all five of these muscles. Intracellular recordings made from muscle fibers near the time of hatching (Embryonic Days 18–21 or Postnatal Days 1–2) indicated that the loss of the ability to contract in response to ATP was due to the total loss of responsiveness to ATP. Surgical denervation of the anterior latissimus dorsi and posterior latissimus dorsi was performed in a series of chicks 1–2 days after hatching, and the ability of these muscles to contract in response to ATP was tested 3–10 days after the surgery. Contractions in response to ATP were present in many of the muscles. Thus denervation of muscles in newly hatched chicks led to the reappearance of sensitivity to ATP. The disappearance of ATP responsiveness shortly after muscles become innervated and the reappearance of ATP responsiveness following denervation suggest that the expression of ATP responsiveness is regulated by motor neurons