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PubMed1996

Cytological analysis of fertilization and early embryonic development in incompatible crosses of Drosophila simulans.

Cytoplasmic incompatibility (CI) is a unique form of male sterility found in numerous insect species that harbor a bacterial endosymbiont Wolbachia. CI is characterized by severe reduction in the progeny produced when infected males are crossed to uninfected females. The reduction in progeny correlates with developmental defects that arise during and immediately following fertilization, suggesting that sperm function is disrupted. To investigate the nature of the cellular defects associated with CI, fertilization and early embryonic development were examined in normal and incompatible crosses of Drosophila simulans using anti-sperm, anti-tubulin and anti-chromatin antibodies. Although pleiotropic, defects associated with CI can be classified into five broad categories: (1) sperm defects in the egg; (2) aberrant morphology of the mitotic apparatus; (3) defects in chromatin structure; (4) proliferation of centrosomes in the absence of nuclear division; and (5) loss of mitotic synchrony. Although mitosis and chromosome behavior are severely disrupted in CI crosses during early development, centrosome duplication and migration appear to continue unabated. The available cytological data suggest that the primary defects observed in incompatible crosses are due to defects in chromosome replication/segregation and in associated centrosome/microtubule-based processes.

Animals↗

PubMed1999

Habitat, world geographic range, and embryonic development of hosts explain the prevalence of avian hematozoa at small spatial and phylogenetic scales.

The factors explaining interspecific differences in prevalences of blood parasites in birds are poorly known. We simultaneously assessed 20 social, ecological, life history, and sampling-related variables that could influence hemoparasite prevalences among diurnal birds of prey in Spain. Our results show that multiple factors are responsible for the studied host-parasite association. We confirmed for the first time that prevalence is inversely correlated to the embryonic development period, and thus probably to immune performance, even among closely related birds. Macrohabitat features related to vector availability are also important, prevalences being higher in species breeding in forested habitats. Finally, prevalence is positively correlated with the host's world geographic range. We hypothesize that larger geographic ranges offered more opportunities for host-vector-hemoparasite associations to become established. The results from our multivariate analyses differ from those obtained through univariate ones, showing that all potential factors should be assessed jointly when testing any ecological or evolutionary hypothesis dealing with parasites.

Animals↗

PubMed1998

BMP-4 is proteolytically activated by furin and/or PC6 during vertebrate embryonic development.

Bone morphogenetic protein-4 (BMP-4) is a multifunctional developmental regulator. BMP-4 is synthesized as an inactive precursor that is proteolytically activated by cleavage following the amino acid motif -Arg-Ser-Lys-Arg-. Very little is known about processing and secretion of BMPs. The proprotein convertases (PCs) are a family of seven structurally related serine endoproteases, at least one of which, furin, cleaves after the amino acid motif -Arg-X-Arg/Lys-Arg-. To examine potential roles of PCs during embryonic development we have misexpressed a potent protein inhibitor of furin, alpha1-antitrypsin Portland (alpha1-PDX) in early Xenopus embryos. Ectopic expression of alpha1-PDX phenocopies the effect of blocking endogenous BMP activity, leading to dorsalization of mesoderm and direct neural induction. alpha1-PDX-mediated neural induction can be reversed by co-expression of downstream components of the BMP-4 signaling pathway. Thus, alpha1-PDX can block BMP activity upstream of receptor binding, suggesting that it inhibits an endogenous BMP-4 convertase(s). Consistent with this hypothesis, alpha1-PDX prevents cleavage of BMP-4 in an oocyte translation assay. Using an in vitro digestion assay, we demonstrate that four members of the PC family have the ability to cleave BMP-4, but of these, only furin and PC6B are sensitive to alpha1-PDX. These studies provide the first in vivo evidence that furin and/or PC6 proteolytically activate BMP-4 during vertebrate embryogenesis.

Animals↗

PubMed1996

Embryonic development of the enteric nervous system of the grasshopper Schistocerca americana.

The enteric nervous system (ENS) of the grasshopper Schistocerca americana is organized into four ganglia located in the foregut (the dorsal unpaired frontal and hypocerebral ganglia, and the paired ingluvial ganglia), and two plexuses that innervate the foregut and midgut. A dorsomedial recurrent nerve and two lateral esophageal nerves connect the ganglia. The midgut plexus is arranged in four nerves running along the midgut surface. In this study, we have focused on the embryonic development of the grasshopper ENS; we have studied the proliferation pattern, morphogenesis, and some aspects of neuronal differentiation by using a number of specific molecular markers. The grasshopper ENS develops early in embryogenesis (25-30%) from three neurogenic zones (NZs) located on the roof of the stomodeum. These NZs slightly invaginate from an epithelial placode. The expression pattern of specific cell surface proteins and the analysis of the mitotic activity showed that NZs cells delaminate from the epithelium, become neuronal precursors, divide symmetrically, and then actively migrate to their final position in the enteric ganglia or plexuses. The grasshopper enteric ganglia are composed of mixed populations of cells from different NZs. The foregut and midgut plexuses are formed by the dispersal of cells from the developing hypocerebral and ingluvial ganglia. The main ENS nerves are pioneered by axons extending anteriorly from hypocerebral and ingluvial neurons. The insect ENS exhibits an enormous variation in design. Several features of the grasshopper program of neurogenesis and pattern of cell migration are compared to other insects, and some evolutionary implications are discussed.

Animals↗

PubMed1977

Environmental influence on ovulation and embryonic development in Rana pipiens.

Environmental effects on ovulation and embryogenesis in Rana pipiens were assessed using both freshly-captured fall animals and laboratory-conditioned females which had undergone vitellogenesis in the laboratory. Frogs in both categories were divided into two groups. Ovulation was hormonally induced in one group of females prior to cold exposure and in the second group of animals following an 8-week-period at 4 degrees C with an 8L 16D photoperiod. The incidence of both ovulation and normal embryonic development was increased following exposure of the animals to low temperatures and short daylength. Those animals which only partially ovulated prior to cold treatment did not respond to hormone injections following the period of cold exposure. Examination of the ovaries of these females revealed a much greater degree of oocyte resorption than was found in frogs whose initial ovulation was induced only after exposure to cold temperatures. The administration of ovulation-inducing hormones prior to artificial hibernation may thus have initiated a phase of oocyte resorption which progressed even at 4 degrees C. The incidence of ovulation was similar in wild-caught and laboratory-conditioned females, but eggs from the latter showed a much lower percentage of development to Shumway stage 20. This effect may have been related to differences in the environmental factors to whcih the two groups were exposed during oogenesis.

Animals↗

PubMed2005

AP-2alpha selectively regulates fragile X mental retardation-1 gene transcription during embryonic development.

Fragile X syndrome (FXS) is almost always caused by silencing of the FMR1 gene. The defects observed in FXS indicate that the normal FMR1 gene has a range of functions and plays a particularly prominent role during development. However, the mechanisms regulating FMR1 expression in vivo are not known. Here, we have tested the role of the transcription factor AP-2alpha in regulating Fmr1 expression. Chromatin immunoprecipitation showed that AP-2alpha associates with the Fmr1 promoter in vivo. Furthermore, Fmr1 transcript levels are reduced >4-fold in homozygous null AP-2alpha mutant mice at embryonic day 18.5 when compared with normal littermates. Notably, AP-2alpha exhibits a strong gene dosage effect, with heterozygous mice showing approximately 2-fold reduction in Fmr1 levels. Examination of conditional AP-2alpha mutant mice indicates that this transcription factor plays a major role in regulating Fmr1 expression in embryos, but not in adults. We further investigated the role of AP-2alpha in the developmental regulation of Fmr1 expression using the Xenopus animal cap assay. Over-expression of a dominant-negative AP-2alpha in Xenopus embryos led to reduced Fmr1 levels. Moreover, exogenous wild-type AP-2alpha rescued Fmr1 expression in embryos where endogenous AP-2alpha had been suppressed. We conclude that AP-2alpha associates with the Fmr1 promoter in vivo and selectively regulates Fmr1 transcription during embryonic development.

Animals↗

PubMed2002

Expression of the Ste20-like kinase SLK during embryonic development and in the murine adult central nervous system.

Cell growth and terminal differentiation are controlled by complex signaling cascades that regulate the expression of specific subsets of genes controlling cell fate and morphogenic processes. We have recently cloned and characterized a novel Ste20-like kinase termed SLK (Sabourin et al., Mol. Cel. Biol. 20 (2000) 684). However, the specific function of SLK is poorly understood. To gain further insights into the role of SLK we have characterized its activity, expression and distribution in the CNS during embryonic development and in the adult brain. Although SLK is expressed ubiquitously in adult tissues, our results show that it is expressed preferentially in neuronal lineages during development. We find that SLK is preferentially expressed in the neurons and neuroepithelium of the developing embryo and can be detected at 10.5 and 12.5 days post-coitum (dpc) in the forebrain, midbrain and hindbrain of the developing CNS. At later stages (14.5 dpc), SLK is expressed in the hypothalamus region, all layers of the neural tube, dorsal root ganglion and in the proliferating ependymal layers. Surprisingly, following middle cerebral artery occlusion, SLK expressing neuronal cells are lost and SLK is localized to phagocytic macrophages/microglia. These results suggest a functional role for SLK in early neuronal development as well as in the adult CNS.

Animals↗

PubMed2003

EXT gene family member rib-2 is essential for embryonic development and heparan sulfate biosynthesis in Caenorhabditis elegans.

EXT gene family members including EXT1, EXT2, and EXTL2 are glycosyltransferases required for heparan sulfate biosynthesis. To examine the biological functions of rib-2, a member of the Caenorhabditis elegans EXT gene family, we generated a mutant worm lacking the rib-2 gene using the UV-TMP method followed by sib-selection. Inactivation of rib-2 alleles induced developmental abnormalities in F2 and F3 homozygous worms, while F1 heterozygotes showed a normal morphology. The F2 homozygous progeny generated from the F1 heterozygous hermaphrodites somehow developed to adult stage but exhibited abnormal characteristics such as developmental delay and egg-laying defects. The F3 homozygous progeny from the F2 homozygous hermaphrodites showed early developmental defects and most of the F3 worms stopped developing during the gastrulation stage. Whole-mount staining analysis for heparan sulfate using Toluidine blue (pH 2.5) revealed a defect of heparan sulfate biosynthesis in the F2 homozygotes. The analysis using fluorometric post-column high-performance liquid chromatography also uncovered reduced production of heparan sulfate in the rib-2 mutant. These results indicate that rib-2 is essential for embryonic development and heparan sulfate biosynthesis in C. elegans.

Animals↗

PubMed1985

mRNA usage during Drosophila melanogaster embryonic development. Analysis of nine cloned DNA segments.

A set of nine phage lambda clones containing inserts from Drosophila melanogaster which are complementary to cDNA made from oocyte poly(A)+ RNA were selected from a larger group. These cloned elements code for a range of middle abundant RNA sequences which show no appreciable change in abundance during Drosophila embryogenesis. Seven of the nine clones are complementary to two oocyte RNAs, one to three RNAs and one to four RNAs. This study describes the changes that occur in these RNAs during embryonic development in the polysomal and non-polysomal fraction, and in the poly(A)+ RNA and poly(A)- RNA fraction. In all nine of these clones, greater than 70% of the complementary RNA is found in the polysomal region of a sucrose gradient. This proportion increases somewhat during development. Specific changes have been found during development in the proportion of RNA that is poly(A)+. Depending to the cloned sequence, this proportion may increase, decrease, or remain unchanged. For those clones that show a change, most of this change occurs between 8 and 19 h of development. Our data suggest, furthermore, the presence of a class of non-adenylated RNA being utilized during embryogenesis.

Animals↗

PubMed1998

Active brood care in an amphipod: influences of embryonic development, temperature and oxygen.

Female amphipods (Crustacea) carry their fertilized eggs in an external brood pouch until the fully formed juveniles emerge (passive brood care). They may also direct specific maternal activities towards the brood (active brood care). We show that Crangonyx pseudogracilis, which typically populates fresh-waters subject to wide fluctuations in temperature and dissolved oxygen, engages in a highly responsive form of active brood care. This involves a flexing motion by the female that expands the brood pouch and increases the suspension of the eggs in the surrounding medium, accompanied by ventilation of the brood pouch and the 'cycling' of eggs therein. Females also selectively eject nonviable eggs from their broods. We investigated the expression of this brood care behaviour in relation to intrinsic and extrinsic factors relevant to the development of broods. The time spent by females in this behaviour initially increased as embryos developed, but decreased once advanced embryos began to self-ventilate and to have a heart pulse. In addition, both increased temperature and decreased oxygen concentration resulted in increased levels of brooding behaviour. We thus propose that this behaviour functions to ameliorate the microclimate of the brood pouch and serves the changing metabolic demand of the brood, as influenced by the interaction of embryonic development with temperature/dissolved oxygen regime. In addition, this behaviour may be a key adaptation facilitating the success of this North American species as an invader of disturbed and polluted freshwaters in Europe and elsewhere. Evidence is emerging that other amphipods associated with harsh environmental conditions also show such active maternal brood care. Copyright 1998 The Association for the Study of Animal Behaviour.

Journal Article↗

PubMed1977

Embryonic development in house fly eggs fertilized with genetically deficient sperm.

In house flies, Musca domestica L., eggs fertilized with sperm that have chromosome deficiencies and duplications do not hatch, but develop to a stage where a fully differentiated, prehatch larva is formed. Fifteen different chromosome translocations involving most of the 10 arms of the 5 autosomes were studied by crossing male translocation heterozygotes to normal females. Egg hatch was reduced to 36-66% depending on the translocation used. Eggs that did not hatch after 24 hours were fixed, stained, and examined for stage of development. Several translocations involving the right arm of chromosome 4 indicate that the region closest to the centromere contains genes that affect the process of syngamy or early cleavage divisions, but do not reduce the ability of the sperm to compete for egg fertilization. Approximately 70% of the autosomal genes can be absent from sperm (not simultaneously but in different crosses) without inhibiting embryonic development.

Animals↗

PubMed2005

Expression and transient nuclear translocation of proprotein convertase 1 (PC1) during mouse preimplantation embryonic development.

Preimplantation embryos express a number of hormones, neuropeptides, and membrane receptors known to derive from proteolytic activation of their precursors by the seven-member family of subtilisin-like, calcium-dependent serine proteinases known as proprotein convertases (PCs). The goal of this study was to determine the pattern of PC expression in mouse preimplantation embryos. Transcripts for all PCs, except PC2, were detected by reverse transcription-polymerase chain reaction (RT-PCR) in unfertilized and fertilized eggs. Furin, PACE4, PC1, and PC7 transcripts remained present at subsequent stages of preimplantation embryonic development, whereas the levels of transcripts for PC4 and PC5 gradually disappeared after the 2-cell stage. Proprotein convertase 1 (PC1) expression was further examined at the protein level. Immunoblotting revealed the presence of the zymogen and mature forms of this enzyme in eggs and embryos. Immunofluorescence laser confocal microscopy showed PC1-specific staining throughout the cytoplasm of unfertilized eggs. After fertilization, surprisingly, the staining was concentrated in pronuclei. It relocated to the cytoplasm at postzygotic stages and was particularly strong at junctions between blastomeres. The nuclear translocation of PC1 in fertilized eggs is probably mediated by its prodomain. Indeed, when transduced in human colon carcinoma LoVo cells, a mutant proPC1 incapable of cleaving off its prodomain was shown to accumulate in the nucleus. Furthermore, when N-terminally fused to green fluorescent protein, this domain was able to direct the reporter protein to the nucleus of these cells. Collectively, these data establish that eggs and preimplantation embryos express various PCs necessary for proteolytic activation of precursors of hormones and growth factors. They also raise the possibility of a nuclear function for PC1 during zygote formation.

Active Transport, Cell Nucleus↗

PubMed1978

Preferential X inactivation in human placenta membranes: is the paternal X inactive in early embryonic development of female mammals?

In placenta membranes of newborn girls carrying electrophoretically distinguishable G6PD alleles, the maternally derived isozyme is expressed preferentially. This phenomenon cannot be explained by allelic differences in enzyme activity or by somatic selection directed against cells with particular G6PD phenotypes. Instead, it may be that in this tissue X inactivation is nonrandom. Preferential expression of the maternal X chromosome, as has been shown in marsupials and in extraembryonic membranes of rodents and now in man, may reflect the state of activity of the X chromosomes in the early stages of female embryonic development.

Female↗

PubMed1995

Oxygen availability and embryonic development in sand snail (Polinices sordidus) egg masses

The oxygen transport physiology of sand snail Polinices sordidus egg masses was investigated using oxygen microelectrodes and open-flow respirometry. P. sordidus eggs are laid in a jelly matrix that rapidly absorbs water and swells into a horseshoe-shaped sausage. The average diameter of these sausages is 37 mm. Eggs are enclosed in capsules that are distributed throughout the jelly matrix, but 65 % of the eggs are located within 3 mm of the outer surface. There is no circulatory or canal system within the matrix so all gas exchange between developing embryos and the environment must occur by diffusion through the jelly matrix. Oxygen tension in the outer layer remains moderately high (PO2>10 kPa) throughout incubation but decreases rapidly in more centrally located regions, so that by day 4 embryos in this region are exposed to extremely hypoxic conditions (PO2<1 kPa). This hypoxia limits oxygen consumption of embryos to low levels and appears to slow embryonic development or even to arrest it. From day 4 onwards, the central region gradually become less hypoxic because the hatching of peripherally located embryos causes the outer layers of the jelly matrix to disintegrate and thus reduces the diffusion distance for oxygen between the centrally located embryos and the surrounding sea water. As the oxygen tension rises, development accelerates and the embryos eventually hatch as viable veligers, apparently unharmed by their prolonged exposure to hypoxia.

Journal Article↗

PubMed1992

Dynamic expression of the cell adhesion molecule fasciclin I during embryonic development in Drosophila.

A number of different cell surface glycoproteins expressed in the central nervous system (CNS) have been identified in insects and shown to mediate cell adhesion in tissue culture systems. The fasciclin I protein is expressed on a subset of CNS axon pathways in both grasshopper and Drosophila. It consists of four homologous 150-amino acid domains which are unrelated to other sequences in the current databases, and is tethered to the cell surface by a glycosyl-phosphatidylinositol linkage. In this paper we examine in detail the expression of fasciclin I mRNA and protein during Drosophila embryonic development. We find that fasciclin I is expressed in several distinct patterns at different stages of development. In blastoderm embryos it is briefly localized in a graded pattern. During the germ band extended period its expression evolves through two distinct phases. Fasciclin I mRNA and protein are initially localized in a 14-stripe pattern which corresponds to segmentally repeated patches of neuroepithelial cells and neuroblasts. Expression then becomes confined to CNS and peripheral sensory (PNS) neurons. Fasciclin I is expressed on all PNS neurons, and this expression is stably maintained for several hours. In the CNS, fasciclin I is initially expressed on all commissural axons, but then becomes restricted to specific axon bundles. The early commissural expression pattern is not observed in grasshopper embryos, but the later bundle-specific pattern is very similar to that seen in grasshopper. The existence of an initial phase of expression on all commissural bundles helps to explain the loss-of-commissures phenotype of embryos lacking expression of both fasciclin I and of the D-abl tyrosine kinase. Fasciclin I is also expressed in several nonneural tissues in the embryo.

Animals↗

PubMed1996

Imaginal cell-specific accumulation of the multicatalytic proteinase complex (proteasome) during post-embryonic development in the tobacco hornworm, Manduca sexta.

The multicatalytic proteinase complex is a multi-subunit, high molecular weight proteinase present in the nucleus and cytoplasm of eukaryotic cells. This catalytic complex is involved in diverse cellular functions as part of the ubiquitin proteolysis system, including non-lysosomal proteolysis, antigen presentation, cell cycle progression, and cell proliferation, and in the programmed death of intersegmental muscles after adult eclosion in the tobacco hornworm moth, Manduca sexta. We have investigated the distribution of the multicatalytic proteinase complex in the central nervous system of this moth. At all stages of post-embryonic development, most cell types exhibited consistent, low levels of cytoplasmic and nuclear immunoreactivity for the multicatalytic proteinase complex. High levels of cell-specific accumulation of the complex were, however, demonstrated in abdominal neurosecretory cells and in imaginal cells in the larval brain, the larval segmental ganglia, and the developing wing discs. Imaginal cells exhibited intense immunoreactivity for the multicatalytic proteinase complex only until the onset of terminal differentiation. Intersegmental muscles undergoing programmed cell death exhibited intense cytoplasmic immunoreactivity for the multicatalytic proteinase, while persisting flight muscles and dying neurons were characterized by basal levels of staining. These staining patterns suggest that the multicatalytic proteinase of Manduca sexta serves multiple functions and is associated with the period of developmental arrest displayed by imaginal cells prior to metamorphosis.

Animals↗

PubMed2002

Changes in mRNA expression of nicotinic acetylcholine receptor subunits during embryonic development of mouse masseter muscle.

Nicotinic acetylcholine receptors (nAChRs) switch from the embryonic-type (alpha 2 beta gamma delta subunits) to the adult-type (alpha 2 beta epsilon delta subunits), and disappear besides the neuromuscular junctions with the development of trunk and limb skeletal muscles. However, little is known about this process during the embryonic development of masseter muscle. To identify the time course of the nAChR transition from embryonic day (E) 11 to the newborn stage in mouse masseter muscle, we analyzed the expression level of delta, epsilon, and gamma subunit mRNAs by competitive polymerase chain reaction in combination with reverse transcription as well as distribution of delta subunit protein by immunohistochemistry. The nAChR delta subunit mRNA was initially detected at E11, showed an approximately 25-fold increase (p < 0.0001) between E11 and E17, and plateaued thereafter until the newborn stage. Immunostaining for delta subunit was observed in the whole portions of masseter myofibers at E17 and birth, suggesting that the nAChR elimination does not begin even at the newborn stage. The epsilon subunit mRNA initially appeared at E17, and increased in quantity by 144% (p < 0.0001) up to the newborn stage. The quantity of gamma subunit mRNA increased by approximately 240% (p < 0.0001) between E11 and E17, and then decreased by 22% (p < 0.05) from E17 value at the newborn stage. The beginning of the expression of the epsilon subunit mRNA was coincident with the beginning of the decrease in the quantity of the gamma subunit mRNA, suggesting that the nAChR subunit switch begins at E17.

Animals↗

PubMed1998

The expression of tenascin-C along the lamprey olfactory pathway during embryonic development and following axotomy-induced replacement of the olfactory receptor neurons.

Extracellular guidance molecules affect the pathway of growing axons by both attractive and repulsive interactions. Tenascin-C, a glycoprotein of the extracellular matrix, is localized along developing axonal pathways where it may function by repulsion, restricting axons within specific boundaries. The lamprey olfactory pathway offers an advantageous model for studying the role of extracellular matrix proteins in axon guidance because the entire pathway is readily seen in horizontal sections and because lesioning the olfactory nerve will induce the system into a new phase of coordinated neurogenesis and axon outgrowth. Although tenascin-C expression was absent during embryonic development, olfactory nerve fascicles contained tenascin-C-immunoreactivity (IR) during the larval stage. During retrograde degeneration, the fascicles lost tenascin-C-IR. Diffuse unfasciculated axonal processes extending from the olfactory epithelium did not express tenascin-C-IR; however, acetylated tubulin and GAP-43-IR was present, indicating axonal outgrowth. When the newly extended axons of olfactory receptor neurons converged to form fascicles, tenascin-C-IR was evident within the fascicular boundaries. The absence of tenascin-C expression when axonal process were short and diffuse, and its return when axons coalesced within fascicles, supports the view that tenascin-C functions as a boundary molecule in the olfactory pathway.

Animals↗