pinecone/core-2020-05-10-deduplication huggingface dataset:
Genetic Analysis of Tissue Interactions Required for Otic Placode Induction in the Zebrafish

AbstractDevelopment of the vertebrate inner ear begins during gastrulation with induction of the otic placode. Several embryonic tissues, including cephalic mesendoderm, notochord, and hindbrain, have been implicated as potential sources of otic-inducing signals. However, the relative contributions of these tissues have not been determined, nor have any genes affecting placode induction been identified. To address these issues, we analyzed otic placode induction in zebrafish mutants that are deficient in prospective otic-inducing tissues. Otic development was monitored by examining mutant embryos for morphological changes and, in some cases, by visualizing expression patterns ofdlx-3orpax-2.1in preotic cells several hours before otic placode formation. Incyclops(cyc−) mutants, which develop with a partial deficiency of prechordal mesendoderm, otic induction is delayed by up to 1 h. Inone-eyed pinhead(oep−) mutants, which are more completely deficient in prechordal mesendoderm, otic induction is delayed by 1.5 h, and morphology of the otic vesicles is abnormal. Expression of marker genes in other regions of the neural plate is normal, suggesting that ablation of prechordal mesendoderm selectively inhibits otic induction. In contrast, the timing and morphology of otic development is not affected by mutations inno tail(ntl) orfloating head(flh), which prevent notochord differentiation. Similarly, a mutation invalentino(val), which blocks early differentiation of rhombomeres 5 and 6 in the hindbrain, does not delay otic induction, although subsequent patterning of the otic vesicle is impaired. To test whether inductive signals from one tissue can compensate for loss of another, we generated double or triple mutants with various combinations of the above mutations. In none of the multiple mutants do theflhorvalmutations exacerbate delays in placode induction, althoughvaldoes contribute additively to defects in subsequent patterning of the otic vesicle. In contrast, mutants homozygous for bothoepandntl, which interact synergistically to disrupt differentiation of cephalic and axial mesendoderm, show a delay in otic development of about 3 h. These data suggest that cephalic mesendoderm, including prechordal mesendoderm and anterior paraxial mesendoderm, provides the first otic-inducing signals during gastrulation, whereas chordamesoderm plays no discernible role in this process. Becauseval−mutants are deficient for only a portion of the hindbrain, we cannot rule out a role for that tissue in otic placode induction. However, if the hindbrain does provide otic-inducing signals, they apparently differ quantitatively or qualitatively from the signals required for vesicle patterning, asvaldisrupts only the latter
RNA-Mediated Interference of acdc25Homolog inCaenorhabditis elegansResults in Defects in the Embryonic Cortical Membrane, Meiosis, and Mitosis

AbstractThe CDC25 dual-specificity phosphatase family has been shown to play a key role in cell cycle regulation. The phosphatase activity of CDC25 drives the cell cycle by removing inhibitory phosphates from cyclin-dependent kinase/cyclin complexes. Although the regulation of CDC25 phosphatase activity has been elucidated both biochemically and genetically in other systems, the role of this enzyme during development is not well understood. To examine the expression pattern and function of CDC25 inCaenorhabditis elegans, we characterized acdc25homolog,cdc-25.1, during early embryonic development. The CDC-25.1 protein localizes to oocytes, embryonic nuclei, and embryonic cortical membranes. When the expression of CDC-25.1 was disrupted by RNA-mediated interference, the anterior cortical membrane of fertilized eggs became very fluid during meiosis and subsequent mitotic cell cycles. Mispositioning of the meiotic spindle, defects in polar body extrusion and chromosome segregation, and abnormal cleavage furrows were also observed. We conclude that CDC-25.1 is required for a very early developmental process—the proper completion of meiosis prior to embryogenesis
The Dominant MutationGlazedIs a Gain-of-Function Allele ofwinglessThat, Similar to Loss of APC, Interferes with Normal Eye Development

AbstractDominant mutations have served as invaluable tools forDrosophilageneticists. Here we analyze the dominant eye mutationGlazed(Gla) that was described by T. H. Morgan more than 50 years ago. We show thatGlacauses the loss of photoreceptor cells during pupal stages, in a process reminiscent of apoptosis, with a concomitant overproduction of eye pigment. This phenotype is very similar to that caused by the loss of D-APC, a negative regulator of Wingless (Wg) signal transduction. Genetic analyses reveal however that theGlagain-of-function phenotype can be reverted to wild-type. By generating a P-element-induced revertant ofGlawe demonstrate thatGlais allelic towg. The molecular lesion inGlaindicates that the insertion of arooretrotransposon leads to ectopic expression ofwgduring pupal stages. We show that theGlaphenotype is similar to that caused by ectopic expression of Wg driven by thesevenless(sev) enhancer. In both cases Wg exerts its effect, at least in part, by negatively regulating the expression of the Pax2 homologsparkling(spa).Glarepresents not only the first dominant allele ofwg, but it may also be the first allele ever described forwg
Processing of the Sperm Protein Sp17 during the Acrosome Reaction and Characterization as a Calmodulin Binding Protein

AbstractIn this study we have demonstrated that the native rabbit sperm protein, Sp17, is a 22- to 24-kDa triplet of proteins in washed ejaculated rabbit spermatozoa and is unaffected by capacitation. However, during the acrosome reaction, Sp17 is processed from a 22- to 24-kDa triplet of proteins to a triplet of proteins at 17–19 kDa by the removal of amino acids from the C-terminal. Recombinant rabbit Sp17 (rRSp17) can also be proteolytically processed by acrosome-reacted spermatozoa in a similar manner. Protease inhibitors prevent the proteolytic processing of Sp17. Both forms of native Sp17 remain associated with acrosome-reacted spermatozoa and are solubilized by ionic detergents. Previously, sequence analysis of Sp17 revealed that Sp17 amino acids 108–137 were 52% identical to the calmodulin binding domain of neuromodulin and contained an IQ motif found in other calmodulin binding proteins. In this study, a truncated recombinant Sp17, rRSp17CB, which lacks amino acids 118–146, including the potential calmodulin binding site, was made. Recombinant rabbit Sp17, but not rRSp17CB, binds to calmodulin in the presence of Ca2+or EDTA, under reduced or nonreduced conditions in biotinylated-calmodulin overlay assays. In DSS crosslinker experiments, calmodulin bound to rRSp17 in a 1:1 ratio but not to rRSp17CB. Additionally, biotinylated rRSp17 interacts with native sperm calmodulin. We propose that the processing of native Sp17, by removing a C-terminal fragment during the acrosome reaction, might be a mechanism to regulate the calmodulin binding activity of Sp17 and provide calmodulin at specific sites after the acrosome reaction
The Synthesis of a Small Heat Shock/α-Crystallin Protein inArtemiaand Its Relationship to Stress Tolerance during Development

AbstractFertilized oocytes of the brine shrimpArtemia franciscanaundergo either ovoviviparous or oviparous development, yielding free-swimming larvae (nauplii) or encysted gastrulae (cysts), respectively. Encystment is followed by diapause, wherein metabolism is greatly reduced; the resulting cysts are very resistant to extreme stress, including desiccation and long-term anoxia. The synthesis of p26, a small heat shock/α-crystallin protein produced only in oviparously developingArtemia,is shown in this paper to be transcriptionally regulated. A p26 mRNA of about 0.7 kb was detected on Northern blots in the second day after oocyte fertilization. It peaked as embryos encysted and declined rapidly when activated cysts resumed development. The appearance of p26 protein, as indicated by immunoprobing of Western blots, followed mRNA by 1 day; it also increased as encystment occurred but remained constant during postgastrula development of cysts. However, p26 underwent a marked reduction during emergence of nauplii and could not be detected in cell-free extracts of second-instar larvae. p26 entered nuclei of encysting embryos soon after synthesis and was localized therein as late as instar II, when it was restricted to a small set of salt gland nuclei. First-instar larvae derived from cysts were more thermotolerant than larvae that had developed ovoviviparously, but synthesis of p26 was not induced by heat under the experimental conditions employed. Additionally, transformed bacteria synthesizing p26 were more thermotolerant than bacteria that lacked the protein. The results support the proposal that p26, a developmentally regulated protein synthesized during embryo encystment, has chaperone activityin vivoand protects the proteins of encystedArtemiafrom stress-induced denaturation
WNT Signaling in the Control of Hair Growth and Structure

AbstractCharacterization of the molecular pathways controlling differentiation and proliferation in mammalian hair follicles is central to our understanding of the regulation of normal hair growth, the basis of hereditary hair loss diseases, and the origin of follicle-based tumors. We demonstrate that the proto-oncogeneWnt3,which encodes a secreted paracrine signaling molecule, is expressed in developing and mature hair follicles and that its overexpression in transgenic mouse skin causes a short-hair phenotype due to altered differentiation of hair shaft precursor cells, and cyclical balding resulting from hair shaft structural defects and associated with an abnormal profile of protein expression in the hair shaft. A putative effector molecule for WNT3 signaling, the cytoplasmic protein Dishevelled 2 (DVL2), is normally present at high levels in a subset of cells in the outer root sheath and in precursor cells of the hair shaft cortex and cuticle which lie immediately adjacent toWnt3-expressing cells. Overexpression ofDvl2in the outer root sheath mimics the short-hair phenotype produced by overexpression ofWnt3,supporting the hypothesis thatWnt3andDvl2have the potential to act in the same pathway in the regulation of hair growth. These experiments demonstrate a previously unrecognized role for WNT signaling in the control of hair growth and structure, as well as presenting the first example of a mammalian phenotype resulting from overexpression of aDvlgene and providing an accessiblein vivosystem for analysis of mammalian WNT signaling pathways
Cloning and Functional Studies of a Novel Gene Aberrantly Expressed in RB-Deficient Embryos

AbstractThe tumor suppressor RB regulates diverse cellular processes such as G1/S transition, cell differentiation, and cell survival. Indeed,Rb-knockout mice exhibit phenotypes including ectopic mitosis, defective differentiation, and extensive apoptosis in the neurons. Using differential display, a novel gene,Rig-1,was isolated based on its elevated expression in the hindbrain and spinal cord ofRb-knockout embryos. The longest open reading frame ofRig-1encoded a polypeptide that consists of a putative extracellular segment with five immunoglobulin-like domains and three fibronectin III-like domains, a putative transmembrane domain, and a distinct intracellular segment. The Rig-1 sequence was 40% identical to the recently identified roundabout protein. Consistent with the predicted transmembrane nature of the protein, Rig-1 protein was present in the membranous fraction. Antisera raised against the putative extracellular and intracellular segments of Rig-1 reacted with an ∼210-kDa protein in mouse embryonic CNS. Rig-1 mRNA was transiently expressed in the embryonic hindbrain and spinal cord. Elevated levels of Rig-1 mRNA and protein were found inRb−/−embryos. Ectopic expression of a transmembrane form of Rig-1, but not the secreted form, promoted neuronal cell entrance to S phase and repressed the expression of a marker of differentiated neuron, Tα1 tubulin. ThusRig-1,a possible distant relative ofroundabout,may mediate some of the pleiotropic roles of RB in the developing neurons
Disrupted Retinal Development in the Embryonic Belly Spot and Tail Mutant Mouse

AbstractTheBelly spot and tail(Bst) semidominant mutation, mapped to mouse Chromosome 16, leads to developmental defects of the eye, skeleton, and coat pigmentation. In the eye, the mutant phenotype is characterized by the presence of retinal colobomas, a paucity of retinal ganglion cells, and axon misrouting. The severity of defects in theBst/+ retina is variable among individuals and is often asymmetric. In order to determine the role of theBstlocus during retinal morphogenesis, we searched for the earliest observable defects in the developing eye. We examined the retinas ofBst/+ and +/+ littermates from embryonic day 9.5 (E9.5) through E13.5 and measured retinal size, cell density, cell death, mitotic index, and cell birth index. We have found that development of theBst/+ retina is notably dilatory by as early as E10.5. The affected retinas are smaller than their wildtype counterparts, and optic fissure fusion is delayed. In the mutant, there is a marked lag in the exit of retinal cells from the mitotic cycle, even though there are no observable differences in the rate of cellular proliferation or cell death between the two groups. We hypothesize thatBstregulates retinal cell differentiation and that variability of structural defects in the mutant, such as those affecting optic fissure fusion, is a reflection of the extent of developmental delay brought about by theBstmutation
Bone Morphogenetic Protein Antagonism of Spemann's Organizer Is Independent of Wnt Signaling

AbstractTheXenopushomeobox genetwinis involved in the Wnt-mediated induction of Spemann's organizer. Additionally, several lines of evidence indicate that bone morphogenetic proteins (BMPs) play a role in repressing the formation of the organizer by antagonizing the expression of genes involved in organizer establishment. In order to determine at what level BMPs exert their effect, we measured the activity of different genes expressed within the organizer region. We report that BMP signaling can antagonize the induction of the dorsal-specific genegoosecoidbut is unable to affect Wnt signaling at the level oftwin.These results suggest that the antagonistic activities of BMPs in organizer formation occur postzygotically, independent oftwinregulation, and that Wnt-like dorsal determinant signaling pathways do not crosstalk with BMPs
Vezf1:A Zn Finger Transcription Factor Restricted to Endothelial Cells and Their Precursors

AbstractUsing retroviral entrapment vectors, we identified a novel mouse gene whose expression is restricted to vascular endothelial cells and their precursors in the yolk sac blood islands. A 3.68-kb cDNA corresponding to the endogenous transcript was isolated using genomic DNA flanking the entrapment vector insertion as a probe. We have named this geneVezf1for vascular endothelial zinc finger 1.Vezf1encodes a protein with a predicted molecular mass of 56 kDa and that contains six putative zinc finger domains and shows high homology to a previously identified human gene, DB1, that is believed to be involved in regulating expression of cytokine genes such as interleukin-3.In situhybridization analysis revealed the onset of expression in advanced primitive streak-stage embryos being located in the extraembryonic mesodermal component of the visceral yolk sac and in the anteriormost mesoderm of the embryo proper. During head-fold and somite stages, expression was restricted to vascular endothelial cells that arise during both vasculogenesis and angiogenesis.Vezf1-related sequences were found to be highly conserved among higher vertebrate species that have acquired extraembryonic yolk sac membranes during evolution. TheVezf1locus mapped to the proximal part of mouse chromosome 2, a region which has homology to human chromosome 9q.Vezf1expression correlates temporally and spatially with the early differentiation of angioblasts into the endothelial cell lineage and the proliferation of endothelial cells of the embryonic vascular system. Thus,Vezf1may play an important role in the endothelial lineage determination and may have an additional role during later stages of embryonic vasculogenesis and angiogenesis
Identification of PLCγ-Dependent and -Independent Events during Fertilization of Sea Urchin Eggs

AbstractAt fertilization, sea urchin eggs undergo a series of activation events, including a Ca2+action potential, Ca2+release from the endoplasmic reticulum, an increase in intracellular pH, sperm pronuclear formation, MAP kinase dephosphorylation, and DNA synthesis. To examine which of these events might be initiated by activation of phospholipase Cγ (PLCγ), which produces the second messengers inositol trisphosphate (IP3) and diacylglycerol, we used recombinant SH2 domains of PLCγ as specific inhibitors. Sea urchin eggs were co-injected with a GST fusion protein composed of the two tandem SH2 domains of bovine PLCγ and (1) Ca2+green dextran to monitor intracellular free Ca2+, (2) BCECF dextran to monitor intracellular pH, (3) Oregon Green dUTP to monitor DNA synthesis, or (4) fluorescein 70-kDa dextran to monitor nuclear envelope formation. Microinjection of the tandem SH2 domains of PLCγ produced a concentration-dependent inhibition of Ca2+release and also inhibited cortical granule exocytosis, cytoplasmic alkalinization, MAP kinase dephosphorylation, DNA synthesis, and cleavage after fertilization. However, the Ca2+action potential, sperm entry, and sperm pronuclear formation were not prevented by injection of the PLCγSH2 domain protein. Microinjection of a control protein, the tandem SH2 domains of the phosphatase SHP2, had no effect on Ca2+release, cortical granule exocytosis, DNA synthesis, or cleavage. Specificity of the inhibitory action of the PLCγSH2 domains was further indicated by the finding that microinjection of PLCγSH2 domains that had been point mutated at a critical arginine did not inhibit Ca release at fertilization. Additionally, Ca2+release in response to microinjection of IP3, cholera toxin, cADP ribose, or cGMP was not inhibited by the PLCγSH2 fusion protein. These results indicate that PLCγ plays a key role in several fertilization events in sea urchin eggs, including Ca2+release and DNA synthesis, but that the action potential, sperm entry, and male pronuclear formation can occur in the absence of PLCγ activation or Ca2+increase
Neuronal Differentiation and Patterning inXenopus: The Role of cdk5 and a Novel Activator Xp35.2

AbstractCdk5, a member of the cyclin-dependent kinase family, has been shown to play an important role in development of the central nervous system in mammals when partnered by its activator p35. Here we describe the cloning and characterization of a novel activator of cdk5 inXenopus,Xp35.2. Xp35.2 is expressed during development initially in the earliest differentiating primary neurons in the neural plate and then later in differentiating neural tissue of the brain. This is in contrast to the previously describedXenopuscdk5 activator Xp35.1 which is expressed over the entire expanse of the neural plate in both proliferating and differentiating cells. Expression of both Xp35.1 and Xp35.2 and activation of cdk5 kinase occur when terminal neural differentiation is induced by neurogenin and neuro D overexpression but not when only early stages of neural differentiation are induced by noggin. Moreover, blocking cdk5 kinase activity specifically results in disruption and reduction of the embryonic eye where cdk5 and its Xp35 activators are expressed. Thus, cdk5/p35 complexes function in aspects of neural differentiation and patterning in the early embryo and particularly in formation of the eye
Aging-Specific Expression ofDrosophila hsp22

Abstracthsp22is among the least abundantly expressedDrosophilaheat shock (hs) genes during both development and heat stress. In contrast,hsp22was found to be the most abundantly expressed hs gene duringDrosophilaaging. During aging,hsp22RNA was induced 60-fold in the head, with somewhat lower level induction in abdomen and thorax. Induction of the other hs gene RNAs was ≤3-fold, except forhsp23,which was induced ∼5-fold in thorax. hsp22 protein was detected using rat anti-hsp22 polyclonal antisera and was induced >150-fold, with particularly abundant expression in eye tissue. Aging-specific induction ofhsp22was reproduced byhsp22:lacZfusion reporter constructs in transgenic flies. Analysis of specific promoter mutations in transgenic flies indicated that functional heat shock response elements are required forhsp22induction during aging. Finally, comparison ofhsp22RNA and protein expression patterns suggests that aging-specific expression ofhsp22is regulated at both the transcriptional and the posttranscriptional levels. Aging-specific induction ofhsp22is discussed with regard to current evolutionary theories of aging
A Novel Follicle-Cell-Dependent Dominant Female Sterile Allele,StarKojak, Alters Receptor Tyrosine Kinase Signaling inDrosophila

AbstractWe describe a new dominant allele,StarKojak,that alters receptor tyrosine kinase signaling in the follicle cells and in the eyes inDrosophila.We isolatedStarKojakin a screen for follicle-cell-dependent dominant female sterile mutations. We show thatStarKojakand revertants ofStarKojakdo not complementStarloss-of-function mutations. We propose thatStarKojakis a novel type of allele of Star that has both dominant gain-of-function phenotypes early in development and dominant loss-of-function phenotypes later in development.Starencodes a putative transmembrane protein that has previously been shown to be a critical component of the epidermal growth factor receptor tyrosine kinase signaling pathway. Early in oogenesis, Star mRNA expression is higher inStarKojakegg chambers than in wild-type egg chambers, consistent with its gain-of-function phenotype. Later in oogenesis, Star mRNA expression is lower inStarKojakfollicle cells than in wild-type follicle cells, consistent with its loss-of-function phenotype. By genetically analyzingStarKojakand its revertants, we present evidence that Star is involved in anterior–posterior axis formation both in the female germline cells and in the somatic follicle cells. We also demonstrate that at least part of the dominant female sterile phenotype ofStarKojakis restricted to the posterior-pole follicle cells. We propose that Star functions by processing pro-Gurken to mature Gurken, which is thereby released in the region between the oocyte and the follicle cells and binds to the epidermal growth factor receptor in the follicle cells
Wingless Signaling Leads to an Asymmetric Response to Decapentaplegic-Dependent Signaling during Sense Organ Patterning on the Notum ofDrosophila melanogaster

AbstractWnt and Decapentaplegic cell signaling pathways act synergistically in their contribution to macrochaete (sense organ) patterning on the notum ofDrosophila melanogaster.The Wingless-signaling pathway was ectopically activated by removing Shaggy activity (the homologue of vertebrate glycogen synthase kinase 3) in mosaics. Proneural activity is asymmetric within the Shaggy-deficient clone of cells and shows a fixed “polarity” with respect to body axis, independent of the precise location of the clone. This asymmetric response indicates the existence in the epithelium of a second signal, which we suggest is Decapentaplegic. Ectopic expression of Decapentaplegic induces extra macrochaetes only in cells which also receive the Wingless signal. Activation of Hedgehog signaling generates a long-range signal which can promote macrochaete formation in the Wingless activity domain. This signal depends upondecapentaplegicfunction. Autonomous activation of the Wingless signal response in cells causes them to attenuate or sequester this signal. Our results suggest a novel patterning mechanism which determines sense organ positioning inDrosophila