Molecular analyses of yolk syncytial layer formation and its role in zebrafish embryogenesis

卵黄合胞体层形成的分子分析及其在斑马鱼胚胎发生中的作用

• 批准号: BB/F010222/1
• 负责人: Tetsuhiro Kudoh
• 金额: $ 45.36万
• 依托单位: UNIVERSITY OF EXETER
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2008
• 资助国家: 英国
• 起止时间: 2008 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FF010222%2F1
• 关键词: Molecular; analyses; yolk; syncytial; layer

Fish embryos consist of two major compartments: the blastoderm in the animal pole and yolk in the vegetal pole. The blastoderm consists of cells that make the fish body, while the yolk is a structure that is important for storage of nutrients and signalling molecules. Between these two compartments, there is a thin layer of tissue called the Yolk Syncytial Layer (YSL). The YSL is a single cell layer that has many nuclei (the status of a single cell with multiple nuclei is called a syncytium). The precursors of YSL cells are normal cells that have a single nucleus. However, at 500-1000 cell stage, when the YSL is forming, these cell membranes collapse, cells fuse and become single cells with multiple nuclei. The YSL separates the blastoderm and yolk, and the role of the YSL is to transfer and metabolise nutrients as well as to send signals for the patterning of the embryo. As the signalling centre for patterning, the YSL sends signals to neighbouring blastoderm cells and guides these cells to become mesendoderm (precursor of muscle, kidney, gonad, blood, digestive system and others) and also to obtain regional character (e.g. the YSL guides cells to become head, trunk or tail). Although the YSL does not exist in mammals, equivalent cells with similar activities do, therefore the study of the YSL would be of general interest to all vertebrate researchers including physicians studying human development. Despite the importance of the YSL in embryonic development, the molecular mechanisms under which the YSL is formed have not yet been investigated. From our previous genetic screening, we isolated a gene (we named it yugo-b) that is specifically expressed in the YSL. When we knocked down the gene function of yugo-b and related gene, yugo-a, we found that YSL formation is specifically disrupted. Therefore this would be the first gene that has been identified as a crucial regulator of YSL formation. By using this gene as an experimental tool, we aim to investigate the molecular mechanisms of syncytium formation by cell fusion occurring in the YSL. Since the zebrafish embryo is transparent, and YSL formation occurs at specific times in a synchronised manner, the YSL is an ideal system to study cell fusion events occurring in embryonic development. By utilising many tools available in the zebrafish system (genetics, genome information, gene-knockdown using morphlino antisense oligonucleotides, labelling cell structures with fluorescent proteins, whole mount in situ hybridisation, etc.) we aim to obtain a range of results providing insight into the processes of formation of the YSL and the different components, structures, mechanisms and molecules involved.

鱼类胚胎由两个主要部分组成：动物极中的胚盘和植物极中的卵黄。胚盘由构成鱼体的细胞组成，而蛋黄是一种重要的结构，用于储存营养物质和信号分子。在这两个隔室之间，有一层薄薄的组织，称为蛋黄合胞体层（YSL）。YSL是具有许多核的单细胞层（具有多个核的单细胞的状态被称为合胞体）。YSL细胞的前体细胞是具有单个核的正常细胞。然而，在500-1000细胞阶段，当YSL形成时，这些细胞膜塌陷，细胞融合并成为具有多个核的单细胞。YSL分离胚盘和蛋黄，YSL的作用是转移和代谢营养物质以及发送胚胎图案化的信号。作为图案化的信号中心，YSL向邻近的胚盘细胞发送信号，并引导这些细胞成为中内胚层（肌肉、肾脏、性腺、血液、消化系统等的前体），并获得区域特征（例如，YSL引导细胞成为头部、躯干或尾部）。虽然YSL不存在于哺乳动物中，但具有类似活性的等效细胞存在，因此YSL的研究将引起所有脊椎动物研究人员的普遍兴趣，包括研究人类发育的医生。尽管YSL在胚胎发育中的重要性，但YSL形成的分子机制尚未研究。从我们以前的遗传筛选中，我们分离出一个基因（我们命名为yugo-b），它在YSL中特异性表达。当我们敲除yugo-b和相关基因yugo-a的基因功能时，我们发现YSL的形成被特异性地破坏。因此，这将是第一个被确定为YSL形成的关键调节因子的基因。通过使用该基因作为实验工具，我们的目的是研究在YSL中发生的细胞融合合胞体形成的分子机制。由于斑马鱼胚胎是透明的，并且YSL的形成以同步的方式在特定的时间发生，因此YSL是研究胚胎发育中发生的细胞融合事件的理想系统。通过利用斑马鱼系统中可用的许多工具（遗传学、基因组信息、使用吗啉代反义寡核苷酸的基因敲除、用荧光蛋白标记细胞结构、整体原位杂交等），我们的目标是获得一系列的结果，提供洞察的过程中形成的YSL和不同的组成部分，结构，机制和分子参与。