STUDIES ON OVARIAN RING CANALS IN DROSOPHILA

这项工作的长期目标是了解称为环形管道的细胞间连接如何促进合胞体发育。环管起源于被细胞骨架蛋白边缘稳定的有丝分裂沟。它们普遍存在于整个动物王国的生殖细胞发育中，也在许多体细胞中发现。然而，在大多数情况下，环管的功能意义知之甚少。由于环管发育错误而导致的细胞事件协调的中断可能会导致细胞行为的严重畸变，例如生长控制的丧失。实验系统是果蝇卵子发生，在此期间，保育细胞通过环形管道将细胞质转移到卵母细胞来喂养生长中的卵母细胞母体成分。通过研究细胞质运输缺陷导致雌性不育的突变果蝇品系，已经鉴定出几种环管成分。在这项资助申请中提出的工作重点是这些蛋白质的功能分析。具体目标是：（1）鉴定在环管形态发生过程中与细丝蛋白（cheerio编码）相互作用的蛋白质;（2）hts基因编码蛋白质的功能分析和hts蛋白质加工的研究;（3）kelch蛋白功能分析和kelch相互作用蛋白的鉴定。果蝇卵室的环管具有基于肌动蛋白的细胞骨架，其维持环管的完整性，并且还允许环管在致癌基因期间直径增长超过10倍。连接细胞骨架的环管质膜的机制是未知的。细丝蛋白是一个很好的候选人提供这样的联系，并提出研究，以确定蛋白质相互作用的细丝蛋白在卵室。这将提供有关从卵裂沟到环道的过渡的信息。hts基因座在卵泡细胞和生殖细胞中都是必需的。在生殖细胞系中，高温超导产物存在于环管和融合体中。目标2中的研究集中于表征融合体行为和环管组装之间的联系。最后，环管的肌动蛋白细胞骨架是不寻常的，因为细丝排列成一个圆圈。Kelch蛋白对于维持环管生长过程中肌动蛋白的包装组织是重要的。在目标3中，提出了确定kelch的生物化学作用的工作。这项工作将包括高分辨率研究肌动蛋白组织在野生型和kelch突变卵室使用冷冻电子显微镜。

The long-term goal of this work is to understand how intercellular junctions called ring canals contribute to instances of syncytial development. Ring canals originate as arrested mitotic cleavage furrows that become stabilized by a rim of cytoskeletal protein. They are prevalent in developing germ cells throughout the animal kingdom and have also been found in a number of somatic cells. However, the functional significance of ring canals is poorly understood in most of these cases. Disruptions in the coordination of cellular events as a result of faulty ring canal development are likely to cause severe aberrations in cell behavior such as loss of growth control. The experimental system is Drosophila oogenesis, during which nurse cells feed growing oocytes maternal components by transferring cytoplasm to the oocyte through ring canals. By studying mutant fly strains in which defective transport of cytoplasm leads to female sterility, several ring canal components have been identified. The work proposed in this grant application is focused on functional analysis of these proteins. The specific aims of the proposal are: (1) identification of proteins that interact with filamin (encoded by cheerio) during ring canal morphogenesis; (2) functional analysis of proteins encoded by the hts locus and studies on hts protein processing; (3) analysis of kelch protein function and identification of kelch interacting proteins. The ring canals of Drosophila egg chambers have an actin-based cytoskeleton that maintains ring canal integrity and also allows ring canals to grow over 10-fold in diameter during oncogenes. The mechanism for linking the cytoskeleton to the plasma membrane of ring canals is unknown. Filamin is a good candidate for providing such a linkage, and studies are proposed to identify proteins that interact with filamin in egg chambers. This should provide information on the transition from cleavage furrow to ring canal. The hts locus is required in both follicle cells and germline cells. In the germline, hts products are found in both ring canals and the fusome. Studies in aim 2 are focused on characterizing the link between fusome behavior and ring canal assembly. Finally, the actin cytoskeleton of ring canals is unusual because the filaments are arrange in a circle. The kelch protein is important for maintaining the packing organization of actin during the growth of ring canals. In aim 3, work is proposed to define the biochemical role of kelch. The work will include high resolution studies of actin organization in wild type and kelch mutant egg chambers using cryoelectron microscopy.