Mechanisms of Development最新文献

The segment-polarity gene engrailed is required for segmentation in the early Drosophila embryo. Loss of Engrailed function results in segmentation defects that vary in severity from pair-rule phenotypes to a lawn phenotype lacking in obvious of segmentation. During segmentation, Engrailed is expressed in stripes with a single segmental periodicity in Drosophila, which is conserved in all arthropods examined so far. To define segments, the segmental stripes of Engrailed induce the segmental stripes of wingless at each parasegmental boundary. However, segmentation functions of orthologs of engrailed in non-Drosophila arthropods have yet to be reported. Here, we analyzed functions of the Tribolium ortholog of engrailed (Tc-engrailed) during embryonic segmentation. Larval cuticles with Tc-engrailed being knocked down had segmentation phenotypes including incomplete segment formation and loss of a group of segments. In agreement with the cuticle segmentation defects, segments developed incompletely and irregularly or did not form in Tribolium germbands where Tc-engrailed was knocked down. Furthermore, knock-down of Tc-engrailed did not properly express the segmental stripes of wingless in Tribolium germbands. Taken together with the conserved expression patterns of Engrailed in arthropod segmentation, our data suggest that Tc-engrailed is required for embryonic segmentation in Tribolium, and the genetic mechanism of Engrailed inducing wingless expression is conserved at least between Drosophila and Tribolium.

In the 1980', vascular endothelial growth factor (VEGF), also known as vascular permeability factor (VPF) was identified and shown to be a key regulator of endothelial cell proliferation and migration, and vascular permeability. The essential role of VEGF and of VEGF receptors (VEGFRs) for the development of the embryonic vascular system has been established via gene disruption studies. This article summarizes the fundamental contributions that have been made in the early days of angiogenesis research to define the involvement of VEGF/VEGFRs in the developing vasculature.

The formation of the vertebrate nervous system depends on the complex interplay of morphogen signaling pathways and cell cycle progression to establish distinct cell fates. The Sonic hedgehog (Shh) signaling pathway is well understood to promote ventral cell fates in the developing spinal cord. A key regulator of Shh signaling is its receptor Patched1 (Ptch1). However, because the Ptch1 null mutation is lethal early in mouse embryogenesis, its role in controlling cell cycle progression, neurogenesis, and axon guidance in the developing spinal cord is not fully understood. An allele of Ptch1 called Wiggable (Ptch1^Wig), which was previously shown to enhance Shh signaling, was used to test its ability to regulate neurogenesis and proliferation in the developing spinal cord. Ptch1^Wig/Wig mutants displayed enhanced ventral proneural gene activation, and aberrant proliferation of the neural tube and floor plate cells, the latter normally being a quiescent population. The expression of the cell cycle regulators p27^Kip1 and p57^Kip2 were expanded in Ptch1^Wig/Wig mutant spinal cords, as was the number of mitotic and S-phase nuclei, suggesting enhanced cell cycle progression. However, Ptch1^Wig/Wig mutants also showed enhanced apoptosis in the ventral embryonic spinal cord, which resulted in thinner spinal cords at later embryonic stages. Commissural axons largely failed to cross the floor plate of Ptch1^Wig/Wig mutant embryos, suggesting enhanced Shh signaling in these mutants led to a dorsal expansion of the chemoattraction front. These findings are consistent with a role of Ptch1 in regulating neurogenesis and proliferation of neural progenitors, and in restricting the influence of Shh signaling in commissural axon guidance to the floor plate.

The zebrafish offers powerful advantages as a model system for examining the growth of the skull vault and the formation of cranial sutures. The zebrafish is well suited for large–scale genetic screens, available in large numbers, and continual advances in genetic engineering facilitate precise modeling of human genetic disorders. Most importantly, zebrafish are continuously accessible for imaging during critical periods of skull formation when both mouse and chick are physically inaccessible. To establish a foundation of information on the dynamics of skull formation, we performed a longitudinal study based on confocal microscopy of individual live transgenic zebrafish. Discrete events occur at stereotyped stages in overall growth, with little variation in timing among individuals. The frontal and parietal bones initiate as small clusters of cells closely associated with cartilage around the perimeter of the skull, prior to metamorphosis and the transition to juvenile fish. Over a period of ~30 days, the frontal and parietal bones grow towards the apex of the skull and meet to begin suture formation. To aid in visualization, we have generated interactive three–dimensional models based on the imaging data, with annotated cartilage and bone elements. We propose a framework to conceptualize development of bones of the skull vault in three phases: initiation in close association with cartilage; rapid planar growth towards the apex of the skull; and finally overlapping to form sutures. Our data provide an important framework for comparing the stages and timing of skull development across model organisms, and also a baseline for the examination of zebrafish mutants affecting skull development. To facilitate these comparative analyses, the raw imaging data and the models are available as an online atlas through the FaceBase consortium (facebase.org).

Important aspects of vertebrate reproduction, such as gametogenesis, involve changes in organs found deep internally and thus not easily studied directly in most living vertebrates due to obscuring pigment and overlying tissues. Transparent lines of zebrafish, especially the Casper double mutant, allow direct observation and analysis of reproductive events in the gonads in vivo. The natural production of fertilized eggs in zebrafish is a complex process involving oogenesis, spermatogenesis, mating behavior, endocrine and neurological processes with inputs from the environment including light, temperature and nutrition. While these factors play important roles, the hypothalamic-pituitary-gonadal axis (HPGA) is central in the regulation of embryo output. Endocrine disrupting compounds (EDCs) include a variety of pollutants often present in the environment. EDCs may have direct effects on the HPGA or indirect effects through toxic action on supporting organs such as the liver or kidney. Estrogenic compounds such as diethylstilbestrol (DES) have been reported to affect reproduction in a variety of species including man. In this study, the effects of DES on reproduction were determined in a novel way by using transparent Casper zebrafish that allow direct visualization of gonad status over time. Changes in gonad status with DES treatment were correlated with effects on secondary sex characteristics (i.e., genital vent size and breeding tubercles) spawning and embryo production. The results suggest that the Casper zebrafish is a useful model for studying dynamics of reproductive events in vertebrate gonads in vivo and for determining effects of EDCs on zebrafish reproduction.

The establishment of planar cell polarity (PCP) in the Drosophila eye requires correct specification of the R3/R4 pair of photoreceptor cells, determined by a Frizzled mediated signaling event that specifies R3 and induces Delta to activate Notch signaling in the neighboring cell, specifying it as R4. Here, we investigated the role of the Notch signaling negative regulator Numb in the specification of R3/R4 fates and PCP establishment in the Drosophila eye. We observed that Numb is transiently upregulated in R3 at the time of R3/R4 specification. This regulation of Numb levels in developing photoreceptors occurs at the post-transcriptional level and is dependent on Dishevelled, an effector of Frizzled signaling, and Lethal Giant Larva. We detected PCP defects in cells homozygous for numb¹⁵, but these defects were due to a loss of function mutation in fat (fat^Q805⁎) being present in the numb¹⁵ chromosome. However, mosaic overexpression of Numb in R4 precursors (only) caused PCP defects and numb loss-of-function alleles had a modifying effect on the defects found in a hypomorphic dishevelled mutation. Our results suggest that Numb levels are upregulated to reinforce the bias of Notch signaling activation in the R3/R4 pair, two post-mitotic cells that are not specified by asymmetric cell division.

The axial skeleton is divided into different regions based on its morphological features. In particular, in birds and mammals, ribs are present only in the thoracic region. The axial skeleton is derived from a series of somites. In the thoracic region of the axial skeleton, descendants of somites coherently penetrate into the somatic mesoderm to form ribs. In regions other than the thoracic, descendants of somites do not penetrate the somatic lateral plate mesoderm. We performed live-cell time-lapse imaging to investigate the difference in the migration of a somite cell after contact with the somatic lateral plate mesoderm obtained from different regions of anterior–posterior axis in vitro on cytophilic narrow paths. We found that a thoracic somite cell continues to migrate after contact with the thoracic somatic lateral plate mesoderm, whereas it ceases migration after contact with the lumbar somatic lateral plate mesoderm. This suggests that cell–cell interaction works as an important guidance cue that regulates migration of somite cells. We surmise that the thoracic somatic lateral plate mesoderm exhibits region-specific competence to allow penetration of somite cells, whereas the lumbosacral somatic lateral plate mesoderm repels somite cells by contact inhibition of locomotion. The differences in the behavior of the somatic lateral plate mesoderm toward somite cells may confirm the distinction between different regions of the axial skeleton.

Paired box (Pax) proteins function as regulators of coordinated development in organogenesis by controlling factors such as cell growth and differentiation necessary to organize multiple cell types into a single, cohesive organ. Previous work has suggested that Pax transcription factors may regulate diverse cell types through participation in inductive cell-to-cell signaling, which has not been well explored. Here we show that EGL-38, a Pax2/5/8 ortholog, coordinates differentiation of the C. elegans egg-laying system through separate autonomous and non-autonomous functions synchronized by the EGF pathway. We find that EGL-38 protein is expressed at the correct times to both participate in and respond to the EGF pathway specifying uterine ventral (uv1) cell fate, and that EGL-38 is required for uv1 expression of nlp-2 and nlp-7, which are both markers of and participants in uv1 identity. Additionally, we have separated uv1 cell placement and gene expression as distinct hallmarks of uv1 identity and specification, with different dependencies on EGL-38. The parallels between EGL-38 participation in cell signaling events and previous Pax studies argue that coordination of signaling and response to an inductive pathway may be a common feature of Pax protein function.

Long-label retention has been used by many to prove Cairns' immortal strand hypothesis and to identify potential stem cells. Here, we describe two strategies using 5-ethynl-2′-deoxyuridine (EdU) to identify and understand the distribution of long-label-retaining mammary epithelial cells during formation of the mouse mammary ductal system. First, EdU was given upon two consecutive days per week during weeks 4 through 10 and analyzed for label retention at 13 weeks of age. Alternatively, EdU was given for 14 consecutive days beginning at 28 days of age and ending at 42 days of age. Analyses were conducted at >91 days of age (13 weeks). Many more LREC were detected following the second labeling method and their distribution among the subsequently developed ducts. This finding indicated that the early-labeled cells that retained their label were distributed into portions of the gland that developed after the ending of EdU treatment (i.e. 42–>91 days). These observations may have important meaning with respect to the previously demonstrated retention of regenerative capacity throughout the mouse mammary gland despite age or reproductive history. These results suggest LREC may represent long-lived progenitor cells that are responsible for mammary gland homeostasis. Additionally, these cells may act as multipotent stem cells capable of mammary gland regeneration upon random fragment transplantation into epithelium-denuded mammary fat pads.