pinecone/core-2020-05-10-deduplication huggingface dataset:
Left/Right Patterning Signals and the Independent Regulation of Different Aspects ofSitusin the Chick Embryo

AbstractRecently, a pathway of genes which are part of a cascade regulating the side on which the heart forms during chick development was characterized (M. Levinet al.,1995,Cell82, 1–20). Here we extend these previous studies, showing that manipulation of at least one member of the cascade,Sonic hedgehog(Shh), can affect thesitusof embryonic rotation and of the gut, in addition to the heart. Bilateral expression ofShh,which is normally found exclusively on the left, does not result in left isomerism (a bilaterally symmetrical embryo having two left sides) nor in a completesitus inversusphenotype. Instead, misexpression ofShhon the right side of the node, which in turn leads to bilateralnodalexpression, produces a heterotaxia-like condition, where different aspects of laterality are determined independently. Heartsitushas previously been shown to be altered by ectopic Shh and activin. However, the most downstream gene identified in the LR pathway,nodal,had not been functionally linked to heart laterality. We show that ectopic (right-sided)nodalexpression is able to affect heartsitus,suggesting that the randomization of heart laterality observed inShhandactivinmisexpression experiments is a result of changes innodalexpression and thatnodalis likely to regulate heartsitusendogenously. The first defined asymmetric signal in the left–right patterning pathway isShh,which is initially expressed throughout Hensen's node but becomes restricted to the left side at stage 4+. It has been hypothesized that the restriction ofShhexpression may be due to repression by an upstream activin-like factor. The involvement of such an activin-like factor on the right side of Hensen's node was suggested because ectopic activin protein is able to repressShhon the left side of the node, as well as to induce ectopic expression of a normally right-sided marker, the activin receptorcAct-RIIa.Here we provide further evidence in favor of this model. We find that a member of this family,Activin βB,is indeed expressed asymmetrically, only on the right side of Hensen's node, at the correct time for it to be the endogenous asymmetric activin signal. Furthermore, we show that application of follistatin-loaded beads eliminates the asymmetry inShhexpression, consistent with an inhibition of an endogenous member of the activin–BMP superfamily. This combined with the previous data on exogenous activin supports the model thatActivin βBfunctions in the chick embryo to initiateShhasymmetry. While these data extend our understanding of the early signals which establish left–right asymmetry, they leave unanswered the interesting question of how the bilateral symmetry of the embryo is initially broken to define a consistent left–right axis. Analysis of spontaneous chick twins suggests that, whatever the molecular mechanism, left–right patterning is unlikely to be due to a blastodermal prepattern but rather is initiated in a streak-autonomous manner
Haploinsufficient Phenotypes inBmp4Heterozygous Null Mice and Modification by Mutations inGli3andAlx4

AbstractBone morphogenetic protein 4(Bmp4), a vertebrate homolog ofDrosophila decapentaplegic(dpp), encodes a signaling protein with multiple functions during embryogenesis. Most mouse embryos homozygous for theBmp4tm1blhnull allele die around the time of gastrulation, with little or no mesoderm. Two independently derivedBmp4tm1mutations were backcrossed onto the C57BL/6 genetic background. Several independently expressed, incompletely penetrant abnormalities were observed in heterozygotes, including cystic kidney, craniofacial malformations, microphthalmia, and preaxial polydactyly of the right hindlimb. In addition, heterozygotes were consistently underrepresented at weaning. These results indicate thatBmp4gene dosage is essential for the normal development of a variety of organs and for neonatal viability. Two mutations that enhance the penetrance and expressivity of the polydactylous phenotype were identified:Gli3XtJ,a deletion mutation involving a gene encoding a zinc-finger protein related toDrosophilacubitus interruptus, andAlx4tm1rwm,a targeted null mutation in a gene encoding a paired class homeoprotein related toDrosophilaaristaless. All doubleBmp4tm1; Gli3XtJheterozygotes have extensive anterior digit abnormalities of both fore- and hindlimbs, while all doubleBmp4tm1; Alx4tm1heterozygotes display ectopic anterior digits only on the hindlimbs. These genetic interactions suggest a model for the multigenic control of anterior digit patterning during vertebrate limb development
Cell Cycle Length Affects Gene Expression and Pattern Formation in Limbs

AbstractThe relationship between growth and pattern specification during development remains elusive. Some molecules known to function as growth factors are also potent agents of pattern formation. This raises the possibility that growth factors could act in pattern formation via an effect on the cell cycle. We have tested the significance of the length of the cell cycle for gene expression and pattern formation in developing chick limb buds by locally slowing the cell cycle. When anterior cell cycles are lengthened by reversible inhibition of DNA replication or by other means, some genes characteristic of the posterior polarizing region are expressed, and digit duplication is observed. Conversely, when posterior cell cycles are slowed, expression of some posterior-specific genes is inhibited, but the pattern is normal. These results indicate that control of the length of the cell cycle could play a primary role in pattern formation by influencing the complement of genes expressed in a particular region of the embryo
Growth-Promoting Interaction of IGF-II with the Insulin Receptor during Mouse Embryonic Development

AbstractGenetic analyses of dwarfing phenotypes resulting from targeted mutagenesis of the genes encoding the insulin-like growth factors (IGF-I and IGF-II) and their cognate type 1 IGF receptor (IGF1R) have demonstrated that this signaling system is a major determinant of mouse embryonic growth. Of the two IGF ligands, IGF-I interacts exclusively with IGF1R, whereas IGF-II recognizes an additional receptor (XR), because the growth retardation of embryos lacking both IGR1R and IGF-II (30% of normal birthweight) is more severe than that manifested in either class of singleIgf1rorIgf2null mutants (45 and 60% of normal, respectively). To determine whether XR is the insulin receptor (IR), we examined embryos nullizygous for bothIgf1randInsr.While the growth of embryos lacking solely IR is affected very mildly and only at the end of gestation, concomitant absence of IGF1R results in a severe growth-deficiency phenotype (30% of normal size at birth) that is first detected at Embryonic Day 13.5 and is also characterized by transient edema, curly tail, generalized organ hypoplasia, including the muscles, developmental delays in ossification, and thin epidermis. TheIgf1r/Insrdouble nullizygotes are phenotypically indistinguishable from double mutants lacking IGF1R and IGF-II and from other double and triple mutants in which all of the IGF ligand/receptor interactions have been eliminated. Therefore, these results provide genetic evidence that the growth-promoting function of IGF-II during mouse embryogenesis is mediated in part by signaling through the insulin receptor
Developmental Regulation of the Laminin α5 Chain Suggests a Role in Epithelial and Endothelial Cell Maturation

AbstractWe have previously shown that mouse and bovine endothelial cells express a novel 400-kDa laminin α chain complexed to β1 and γ1 laminin chains. We describe here purification of this laminin isoform from the conditioned medium of a mouse peripheral lymph node endothelial cell line, SVEC. The laminin α chain was isolated from the laminin complex, subjected to Edman digestion, and the amino acid sequences of the resulting peptides were determined. Amino acid sequence revealed 100% identity to the predicted amino acid sequence of the recently reported laminin α5 gene. A monoclonal antibody to the laminin α5 chain was raised (4G6), allowing investigation of its distribution in embryonic, newborn, and mature mouse tissues. The laminin α5 chain was expressed mainly by epithelial, endothelial, and myogenic cells: In both embryonic and mature tissues the laminin α5 chain was strongly expressed by epithelial cells, the bronchi of the lungs and the developing kidney tubules being the sites of strongest expression. However, laminin α5 was not associated with early stages of epithelial cell development, but rather with epithelial cell maturation. Widespread expression of laminin α5 in endothelial cells was apparent only in tissues of mature mice, its appearance correlating approximately with sexual maturity. During embryogenesis and in newborn tissues, laminin α5 occurred in basement membranes of larger blood vessels only, excluding a role in angiogenic processes. Smooth muscle and skeletal muscle cells were the only other cell types which showed considerable laminin α5 expression, with skeletal muscle exhibiting a developmentally regulated pattern of expression: The laminin α5 chain occurred in skeletal muscle fiber basement membranes early in embryogenesis (E13–E15) but decreased with development, remaining strongly expressed only at the neuromuscular junction. The data show that laminin α5 expression is associated with epithelial and endothelial cell maturation, implicating a role for this laminin chain in the maintenance of differentiated epithelial and endothelial cell phenotype
Developmental Regulation of Cell Interactions in theArabidopsis fiddlehead-1Mutant: A Role for the Epidermal Cell Wall and Cuticle

AbstractAlthough the plant epidermis serves primarily a protective role, during plant development some epidermal cells specialize, becoming competent to interact not only with pollen but also with other epidermal cells. In the former case, these interactions mediate recognition, germination, and pollen growth responses and, in the latter case, result in interorgan fusions which, most commonly, alter floral architecture in ways that are thought to promote reproductive success. In either case, all of the initial signaling events must take place across the cell wall and cuticle. InArabidopsis,mutation of theFIDDLEHEADgene alters the shoot epidermis such that all epidermal cells become competent to participate in both types of interactions. Infdh-1mutants, epidermal cells manifest not only a contact-mediated fusion response but also interact with pollen. Since carpel epidermal derivatives manifest both of these properties, we postulated thatfdh-1epidermal cells were ectopically expressing a carpel-like program. In this report we demonstrate that manifestation of thefdh-1phenotype does not require the product of theAGAMOUSgene, indicating that the phenotype is either independent of the carpel development program or thatfdh-1mutations activate a carpel-specific developmental program downstream of theAGgene. Furthermore, we demonstrate that plants bearing mutations in thefdh-1gene show significant changes in cell wall and cuticular permeability. Biochemical analyses of the lipid composition of the crude cell wall fraction reveal thatfdh-1cell walls differ from wild-type and manifest significant changes in high-molecular-weight lipid peaks. These results suggest that cell wall and cuticular permeability may be important determinants in developmental signaling between interacting cells and implicate lipids as important factors in modulating the selectivity of the permeability barrier presented by the epidermal cell wall and cuticle
MouseOtlx2/RIEGExpression in the Odontogenic Epithelium Precedes Tooth Initiation and Requires Mesenchyme-Derived Signals for Its Maintenance

AbstractThe mouseOtlx2gene is a new member of thepaired-like family of homeobox genes whose human homologue,RIEG,is involved in Rieger syndrome, an autosomal-dominant disorder. One of the cardinal features of Rieger syndrome is dental hypoplasia, indicating thatOtlx2/RIEGactivity is essential for normal tooth development. Here, we analyzed the expression ofOtlx2during mouse tooth development and studied its regulation in dental explants.Otlx2expression distinguishes stomatodeal from other ectoderm as early as Embryonic Day 8.5, well before tooth initiation. Thereafter, its craniofacial expression becomes restricted to the tooth-forming areas and to the epithelial components of molar and incisor primordia. AlthoughOtlx2induction precedes the specification of odontogenic mesenchyme, tissue recombination experiments show that the maintenance of its expression requires signals from the mesenchyme and that dental mesenchyme has the capacity to induce ectopic expression ofOtlx2in nondental epithelium. Finally, we compareOtlx2expression with that of the recently identified homeodomain transcription factorBarx1expressed in molar mesenchyme. Their strictly complementary expression patterns in the epithelial and mesenchymal components suggest that both genes participate in the reciprocal tissue interactions which are a hallmark of odontogenesis
Introduction of Cyclin B Induces Activation of the Maturation-Promoting Factor and Breakdown of Germinal Vesicle in Growing Zebrafish Oocytes Unresponsive to the Maturation-Inducing Hormone

AbstractWhen treated with 17α,20β-dihydroxy-4-pregnen-3-one (17α,20β-DP), a natural maturation-inducing hormone in fishes, fully grown zebrafish oocytes are induced to mature via the activation of the maturation-promoting factor (MPF), which consists of cdc2 (a catalytic subunit) and cyclin B (a regulatory subunit). In contrast, 17α,20β-DP is unable to induce growing (previtellogenic and vitellogenic) oocytes to mature. To know the reason growing oocytes fail to mature upon 17α,20β-DP treatment, we investigated changes in the components of machinery responsible for MPF activation during zebrafish oogenesis. Immunoblotting experiments using monoclonal antibodies against cdc2, cyclin B, and cdk7 (an activator of cdc2) have revealed that the concentrations of cdc2 and cdk7 are almost constant during oogenesis. Cyclin B was present in mature oocytes but absent in growing and fully grown immature oocytes. These results, which are identical to those in goldfish, strongly suggest that cyclin B is synthesized from stored (masked) mRNA after 17α,20β-DP stimulation and that its binding to the preexisting cdc2 allows cdk7 to activate MPF. Microinjection of cyclin B protein induced MPF activation and germinal vesicle breakdown in growing oocytes, as well as in fully grown oocytes, indicating that cdk7 present in growing oocytes is already active. Northern blot analysis revealed the presence of cyclin B mRNA in both previtellogenic and fully grown oocytes. These results indicate that, as in fully grown oocytes, growing oocytes are already equipped with the catalytic subunit of MPF (cdc2) and its activator (cdk7) and that the appearance of the regulatory subunit of MPF (cyclin B) is sufficient for initiating maturation. Therefore, the unresponsiveness of growing oocytes to 17α,20β-DP is attributable to a deficiency in the processes leading to cyclin B synthesis, which include 17α,20β-DP reception on the oocyte surface, subsequent signal transduction pathways, and unmasking the stored cyclin B mRNA
The Two Intracellular Ca2+Release Channels, Ryanodine Receptor and Inositol 1,4,5-Trisphosphate Receptor, Play Different Roles during Fertilization in Ascidians

AbstractFertilization in the ascidians triggers an activation wave of calcium release followed by intracellular calcium oscillations synchronous with periodic membrane potential excursions during the completion of the meiotic cell cycle. Fertilization also causes a fast decrease in the egg plasma membrane depolarization-activated calcium current and a large increase in capacitance thought to represent membrane addition to the egg surface. We have analyzed the temporal and causal relationships between these changes in the eggs ofPhallusia mammillatausing whole-cell patch-clamp recording while simultaneously imaging calcium with fura-2 dextran. We have defined the role of ryanodine receptor (RyR) and InsP3receptor (InsP3R) during fertilization and meiosis by looking at the effects of InsP3, cyclic ADP ribose (cADPR), and ryanodine in perfused oocytes. We show that InsP3(10 μMperfused through the patch pipette) is able to trigger sustained oscillations in intracellular calcium concentration in unfertilized oocytes, resembling those recorded in fertilized egg completing meiosis. In addition the sustained oscillations resulting from InsP3perfusion in unfertilized oocytes are sufficient to cause the emission of both polar bodies. In contrast, ryanodine or cADPR never trigger detectable calcium signal in perfused oocytes. Instead, nanomolar concentrations of ryanodine or cADPR cause a capacitance change, implying a net insertion of membrane to the oocyte surface, and trigger a fast decrease in the depolarization-activated calcium current. Both changes are similar to the changes in conductance and capacitance naturally observed following fertilization. These effects, although not associated with measurable calcium signals, are abolished by coperfusion of the calcium chelator BAPTA. In contrast to ryanodine or cADPR, sustained perfusion of the oocyte with nanomolar concentrations of InsP3causes no capacitance change and a slow and moderate decrease in calcium current. Our observations on inseminated patch-clamped eggs further indicate that membrane insertion, which starts 15–20 sec after the onset of the membrane conductance change at fertilization, can be altered by interfering with the RyR. Our results imply that, in ascidians, as in some mammals, RyR and InsP3R play distinct roles during fertilization
Rap1 Overexpression Reveals That Activated RasD Induces Separable Defects duringDictyosteliumDevelopment

AbstractOne of theDictyostelium rasgenes,rasD,is expressed preferentially in prestalk cells at the slug stage of development and overexpression of this gene containing a G12T activating mutation causes the formation of aberrant multitipped aggregates that are blocked from further development (Reymondet al.,1986,Nature,323, 340–343). The ability of theDictyostelium rap1gene to suppress this abnormal developmental phenotype was investigated. Therap1gene and G12V activated and G10V negative mutant forms of therap1gene were independently linked to therasDpromoter and each construct used to transform M1, aDictyosteliumcell line expressing RasD[G12T]. Transformants of M1 that expressed Rap1 or Rap1[G12V] protein still formed multitipped aggregates, but most tips were able to complete development and form fruiting bodies. Cell lines showing this modified phenotype were designated ME (multitipped escape). Therap1[G10V] construct did not modify the M1 phenotype. These data suggest that overexpression of RasD[G12T] has two effects, the formation of a multitipped aggregate and a block in subsequent differentiation and that the expression of Rap1 or Rap1[G12V] reverses only the latter. Differentiation of ME cells in low density monolayers showed the identical low level of stalk and spore cell formation seen for M1 cells under the same conditions. Thus the cell autonomous defect in monolayer differentiation induced in the M1 strain was not corrected in the ME strain. Cell type-specific gene expression during the development of M1 cells is dramatically altered: prestalk cell-specific gene expression is greatly enhanced, whereas prespore-specific gene expression is almost suppressed (Louiset al.,1997,Mol. Biol. Cell,8, 303–312). During the development of ME cells,ecmA mRNA levels were restored to those seen for Ax3, andtagB mRNA levels were also markedly reduced, although not to Ax3 levels.cotCexpression in ME cells was enhanced severalfold relative to M1, although levels were still lower than those observed during the development of Ax3. The low expression ofcar1mRNA during early development of the M1 strain remained low during the development of ME cells. These data are consistent with the idea that the expression of RasD[G12T] affects two independent and temporally separated events and that only the later defect is reversed byrap1
Characterization of the Sea Urchin Major Vault Protein: A Possible Role for Vault Ribonucleoprotein Particles in Nucleocytoplasmic Transport

AbstractVaults are large ribonucleoprotein particles that have been identified in a wide range of eukaryotic organisms. Although present in thousands of copies per cell, their function remains unknown. In this report, we identify the major vault protein in sea urchins as a 107-kDa polypeptide that copurifies with microtubules and ribosomes. Although initially identified in microtubule preparations, the sea urchin major vault protein is not predominantly microtubule-associatedin vivo.Rather, the sea urchin major vault protein is present throughout the cytoplasm in eggs and embryos and in the nucleus in adult somatic cells. Within the nucleus, the sea urchin major vault protein is concentrated in the region of the nucleolus and to punctate regions of the nuclear envelope. In addition, the vault protein localizes to short linear strings juxtaposed to the exterior of the nucleus and extending outward into the cytoplasm. Based on their copurification and intracellular distribution, vaults may be involved in the nucleocytoplasmic transport of ribosomes and/or mRNA
Skeletal and Cardiac α-Actin Isoforms Exhibit Unanticipated Temporal and Tissue-Specific Gene Expression Patterns in Developing Avian Limbs and Embryos

AbstractThe initial expression of skeletal muscle structural genes typically occurs after myogenic determination factor gene expression. We investigated this temporal relationship via a reverse transcription–polymerase chain reaction (RT–PCR) analysis of skeletal and cardiac α-actin (s- and c-actin) mRNA during chick limb development. c-actin transcripts were first detected at the beginning of muscle cytodifferentiation in stage 24/25 limbs, shortly after the initial appearance of MyoD and myogenin mRNAs, and were not detected in nonmyogenic tissues. In contrast, s-actin mRNA was detected in limbs at stage 15–16, periods when myf5 and MRF4 but not MyoD and myogenin transcripts are present (Lin-Jones, J., and Hauschka, S. D.,Dev. Biol.174, 407–422, 1996). While s-actin mRNA was not detected in E7 neural retina and was at variable levels in stage 9–15 neural tube, significant levels were consistently detected in mesodermal tissues which contribute nonmyogenic cells to the limb: stage 9–12 lateral plate and distal portions of stage 25/26 limbs. s-actin transcripts detected in the earliest limbs could thus be in myogenic and/or nonmyogenic cells. These data indicate that while c-actin expression is activated at the onset of limb muscle cytodifferentiation, s-actin expression occurs much earlier, as well as in some nonmyogenic tissues. Whether the precocious expression of s-actin plays a functional role in muscle development remains to be determined