Morphofunctional transformations of the yolk syncytial layer during zebrafish development

The yolk syncytial layer (YSL) is a provisory extraembryonic structure of teleost fishes and representatives of some other taxa with meroblastic cleavage. The YSL of teleosts is a symplast with polymorphous polyploid nuclei. It is known to perform nutritional, morphogenetic, immune, and, probably, other functions. Data about the YSL organization, functioning and regulation is fragmentary. Although gene expression patterns and other aspects of YSL functioning have been studied in Danio rerio, the morphology of its YSL has not been described in detail. The study of zebrafish YSL structure on sequential developmental stages is necessary to recognize specific features of this important polyfunctional system in this model organism and to extend our knowledge about provisory systems. The thickness of the YSL and the distribution of its nuclei are not uniform on each stage and change during development. During oblong and sphere stages the internal YSL (I‐YSL) is filled with yolk inclusions; interphase yolk syncytial nuclei (YSN) and mitotic asters can be seen. During doming and epiboly the external YSL (E‐YSL) is thicker than I‐YSL. On the subsequent stages the YSL is thickened caudally. The dorsal YSL part is thickened during early segmentation stages and becomes the thinnest YSL region later. The anterior part of the YSL is thin, but enlarges during larval period. The YSN of different size and diverse forms, from regular to lobed, are present and form clusters. The number of irregular‐shaped nuclei increases during development. The YSL thickens in the end of endotrophic and in the course of endo‐exotrophic period, and its cytoplasm contains numerous yolk inclusions. After yolk exhaustion the YSL is flat. As the YSL degrades , the YSN become pycnotic, and the YSL remnant probably is cleared by phagocytes. J. Morphol. 275:206–216, 2014. © 2013 Wiley Periodicals, Inc.     

A role for MKP3 in axial patterning of the zebrafish embryo

Fibroblast growth factors (FGFs) are secreted molecules that can activate the RAS/mitogen-activated protein kinase (MAPK) pathway to serve crucial functions during embryogenesis. Through an in situ hybridization screen for genes with restricted expression patterns during early zebrafish development, we identified a group of genes that exhibit similar expression patterns to FGF genes. We report the characterization of zebrafish MAP kinase phosphatase 3 (MKP3; DUSP6 - Zebrafish Information Network), a member of the FGF synexpression group, showing that it has a crucial role in the specification of axial polarity in the early zebrafish embryo. MKP3 dephosphorylates the activated form of MAPK, inhibiting the RAS/MAPK arm of the FGF signaling pathway. Gain- and loss-of-function studies reveal that MKP3 is required to limit the extent of FGF/RAS/MAPK signaling in the early embryo, and that disturbing this inhibitory pathway disrupts dorsoventral patterning at the onset of gastrulation. The earliest mkp3 expression is restricted to the future dorsal region of the embryo where it is initiated by a maternal beta-catenin signal, but soon after its initiation, mkp3 expression comes under the control of FGF signaling. Thus, mkp3 encodes a feedback attenuator of the FGF pathway, the expression of which is initiated at an early stage so as to ensure correct FGF signaling levels at the time of axial patterning.     

ApoA-II directs morphogenetic movements of zebrafish embryo by preventing chromosome fusion during nuclear division in yolk syncytial layer

The high density lipoprotein (HDL) represents a class of lipid- and protein-containing particles and consists of two major apolipoproteins apoA-I and apoA-II. ApoA-II has been shown to be involved in the pathogenesis of insulin resistance, adiposity, diabetes, and metabolic syndrome. In embryo, apoa2 mRNAs are abundant in the liver, brain, lung, placenta, and in fish yolk syncytial layer (YSL), suggesting that apoa2 may perform a function during embryonic development. Here we find out that apoa2 modulates zebrafish embryonic development by regulating the organization of YSL. Disruption of apoa2 function in zebrafish caused chromosome fusing, which strongly blocked YSL nuclear division, inducing disorders in YSL organization and finally disturbing the embryonic epiboly. Purified native human apoA-II was able specifically to rescue the defects and induced nuclear division in zebrafish embryos and in human HeLa cells. The C terminus of apoA-II was required for the proper chromosome separation during nuclear division of YSL in zebrafish embryos and in human HeLa cells. Our data indicate that organization of YSL is required for blastoderm patterning and morphogenesis and suggest that apolipoprotein apoA-II is a novel factor of nuclear division in YSL involved in the regulation of early zebrafish embryonic morphogenesis and in mammalian cells for proliferation.     

Cell lineage of zebrafish blastomeres. II. Formation of the yolk syncytial layer

Dye coupling and cell lineages of blastomeres that participate in the formation of the yolk syncytial layer (YSL) in the zebrafish Brachydanio rerio have been examined. The YSL is a multinucleate layer of nonyolky cytoplasm underlying the cellular blastoderm at one pole of the giant yolk cell. It forms at the time of the 10th (sometimes 9th) cleavage by a collapse of a set of blastomeres, termed marginal blastomeres, into the yolk cell. Marginal blastomeres possess cytoplasmic bridges to the yolk cell before the YSL forms, and injections of fluorescein-dextran into the cells revealed that bridges between the yolk cell and blastoderm do not persist after this time. Injections of Lucifer yellow revealed that shortly after the YSL forms the yolk cell and blastoderm are dye coupled, presumably by gap junctions, and that this coupling disappears gradually during early gastrulation. Lineage analyses revealed that not all of the progeny of early marginal blastomeres participate in YSL formation. Although some descendants of marginal blastomeres remained on the margin during successive cleavages, neither "compartment" nor "strict lineage" models are sufficient to explain the origin of the YSL. It is proposed that the position of a cell on the blastoderm margin, and not the cell's lineage, determines YSL cell fate.     

Regulation of hhex expression in the yolk syncytial layer, the potential Nieuwkoop center homolog in zebrafish

The Nieuwkoop center is the earliest signaling center during dorsal-ventral pattern formation in amphibian embryos and has been implied to function in induction of the Spemann-Mangold organizer. In zebrafish, Nieuwkoop-center-like activity resides in the dorsal yolk syncytial layer (YSL) at the interface of the vegetal yolk cell and the blastoderm. hex homologs are expressed in the anterior endomesoderm in frogs (Xhex), the anterior visceral endoderm in mice, and the dorsal YSL in zebrafish (hhex). Here, we investigate the control of hhex expression in the YSL. We demonstrate that bozozok (boz) is absolutely required for early hhex expression, while overexpression of boz causes ectopic hhex expression. Activation of Wnt/beta-catenin signaling by LiCl induces hhex expression in wild-type YSL but not in boz mutant embryos, revealing that boz activity is required downstream of Wnt/beta-catenin signaling for hhex expression. Further, we show that the boz-mediated induction of hhex is independent of the Boz-mediated repression of bmp2b. Our data reveal that repressive effects of both Vega1 and Vega2 may be responsible for the exclusion of hhex expression from the ventral and lateral parts of the YSL. In summary, zebrafish hhex appears to be activated by Wnt/beta-catenin in the dorsal YSL, where Boz acts in a permissive way to limit repression of hhex by Vega1 and Vega2.     

斑马鱼胚胎背腹轴建立的分子机制

脊椎动物胚胎发育起始于体轴的建立,是胚胎早期发育过程中最重要的事件之一。Wnt、BMP、Nodal和FGF等多个信号通路协同调控细胞分化和细胞运动,促进胚胎胚层的形成和空间上的分离,调控胚胎背腹轴、前后轴和左右轴线的分化,为胚胎进一步发育勾勒出蓝图。本文主要综述斑马鱼胚胎背腹轴建立的分子机制,包括背部组织中心简介;母源Wnt/β-catenin信号调控背部组织中心形成的分子机制;BMP信号调控背腹轴建立的分子机制。     

Essential role of the yolk syncytial layer for the development of isolated blastoderms from medaka embryos

To investigate a possible role of the yolk syncytial layer (YSL) in the development of the medaka embryo, blastoderms were isolated at different stages of embryogenesis either with or without the layer and were incubated in a culture medium. The blastoderms from cleavage stage embryos (stage 8–9), in which the YSL had not yet formed, developed into an irregular mass of cells. But some of the blastoderms isolated with the YSL from the blastula embryos (stage 10) developed into embryo-like structures with apparent body axes and contained differentiated organs, such as the eye, ear, contractile heart, yolk sac-like sphere and posterior body trunk with notochord. The proportion of such explants increased as the developmental stage proceeded. However, the proportion was much smaller when blastoderms were isolated at the blastula stage without the YSL. These results suggest that the YSL is essential for the development of embryonic structures. At stage 12 (early gastrula), the frequency of formation of such structures was the same among blastoderms with or without the YSL, so that these embryos are apparently committed for pattern formation.     

A major role for zygotic hunchback in patterning the Nasonia embryo

Developmental genetic analysis has shown that embryos of the parasitoid wasp Nasonia vitripennis depend more on zygotic gene products to direct axial patterning than do Drosophila embryos. In Drosophila, anterior axial patterning is largely established by bicoid, a rapidly evolving maternal-effect gene, working with hunchback, which is expressed both maternally and zygotically. Here, we focus on a comparative analysis of Nasonia hunchback function and expression. We find that a lesion in Nasonia hunchback is responsible for the severe zygotic headless mutant phenotype, in which most head structures and the thorax are deleted, as are the three most posterior abdominal segments. This defines a major role for zygotic Nasonia hunchback in anterior patterning, more extensive than the functions described for hunchback in Drosophila or Tribolium. Despite the major zygotic role of Nasonia hunchback, we find that it is strongly expressed maternally, as well as zygotically. Nasonia Hunchback embryonic expression appears to be generally conserved; however, the mRNA expression differs from that of Drosophila hunchback in the early blastoderm. We also find that the maternal hunchback message decays at an earlier developmental stage in Nasonia than in Drosophila, which could reduce the relative influence of maternal products in Nasonia embryos. Finally, we extend the comparisons of Nasonia and Drosophila hunchback mutant phenotypes, and propose that the more severe Nasonia hunchback mutant phenotype may be a consequence of differences in functionally overlapping regulatory circuitry.     

Subdividing the embryo: a role for Notch signaling during germ layer patterning in Xenopus laevis

The development of all vertebrate embryos requires the establishment of a three-dimensional coordinate system in order to pattern embryonic structures and create the complex shape of the adult organism. During the process of gastrulation, the three primary germ layers are created under the guidance of numerous signaling pathways, allowing cells to communicate during development. Cell-cell communication, mediated by receptors of the Notch family, has been shown to be involved in mediating diverse cellular behaviors during development and has been implicated in the regulation of cell fate decisions in both vertebrate and invertebrate organisms. In order to investigate a role for Notch signaling during boundary formation between the mesoderm and endoderm during gastrulation, we manipulated Notch signaling in gastrula stage embryos and examined gene expression in resultant tissues and organs. Our findings demonstrate a much broader role for Notch signaling during germ layer determination than previously reported in a vertebrate organism. Activation of the Notch pathway, specifically in gastrula stage embryos, results in a dramatic decrease in the expression of genes necessary to create many different types of mesodermal tissues while causing a dramatic expansion of endodermal tissue markers. Conversely, temporally controlled suppression of this pathway results in a loss of endodermal cell types and an expansion of molecular markers of mesoderm. Thus, our data are consistent with and significantly extend the implications of prior observations suggesting roles for Notch signaling during germ layer formation and establish an evolutionarily conserved role for Notch signaling in mediating mesoderm-endoderm boundaries during early vertebrate development.     

Structure of the yolk syncytial layer in the larvae of whitefishes: A histological study

The yolk syncytial layer (YSL) is a symplastic provisory system that forms at the early stages of embryogenesis of teleosts. The YSL serves morphogenetic, trophic, and immune functions. Despite its important role in development, data on the structure of YSL is scarce. In the present study, comparative histological research on the features of YSL structure in the development of larvae of four economically important species of Coregonidae (Salmoniformes)—Coregonus peled, Coregonus muksun, Coregonus nasus, and Stenodus leucichthys nelma—is presented. The YSL of the larvae of Coregonidae has a complex, differentiated structure. Functional regionalization of YSL cytoplasm, possibly determined by the specific features of nutrient assimilation, is typical for all aforementioned species. Cytoplasm that encircles a large oil globule appears striated. A division into an inner area, filled with yolk inclusions, and an outer, smoother homogeneous area, can be noted in the cytoplasm surrounding the yolk mass. The oil globule is retained after complete utilization of the yolk mass. The inequality of thickness of YSL along anteroposterior and dorsoventral axes also indicates the functional regionalization. The dorsomedial area of YSL, located under the intestine, is the least thick. Dorsolateral areas are strongly incrassated and envelop the intestine from two sides. Gigantic nuclei of exceptionally complex form are typical for YSL. Specific features apply to the form of YSL and its nuclei. Based on the obtained results, a fundamental similarity in organization of the YSL of larvae of the studied whitefishes can be concluded; however, its specific variations distinguishable on a histological level can be discussed.     

Computational analysis of BMP gradients in dorsal-ventral patterning of the zebrafish embryo

The genetic network controlling early dorsal-ventral (DV) patterning has been extensively studied and modeled in the fruit fly Drosophila. This patterning is driven by signals coming from bone morphogenetic proteins (BMPs), and regulated by interactions of BMPs with secreted factors such as the antagonist short gastrulation (Sog). Experimental studies suggest that the DV patterning of vertebrates is controlled by a similar network of BMPs and antagonists (such as Chordin, a homologue of Sog), but differences exist in how the two systems are organized, and a quantitative comparison of pattern formation in them has not been made. Here, we develop a computational model in three dimensions of the zebrafish embryo and use it to study molecular interactions in the formation of BMP morphogen gradients in early DV patterning. Simulation results are presented on the dynamics BMP gradient formation, the cooperative action of two feedback loops from BMP signaling to BMP and Chordin synthesis, and pattern sensitivity with respect to BMP and Chordin dosage. Computational analysis shows that, unlike the case in Drosophila, synergy of the two feedback loops in the zygotic control of BMP and Chordin expression, along with early initiation of localized Chordin expression, is critical for establishment and maintenance of a stable and appropriate BMP gradient in the zebrafish embryo.