ACTIN-BINDING PROTEINS AND DROSOPHILA EMBRYOS

早期果蝇胚胎正被用来研究蛋白质的作用 它们与肌动蛋白合作，在如此大的 和高度动态的合体细胞。肌动蛋白细丝结构参与 在细胞质的组织和所需的动态事件中 在正常发育过程中。因此，这些研究将提供对 肌动蛋白介导的过程，有助于形成适当的 有组织的细胞胚层作为分化的底物。自.以来 肌动蛋白的结构和过程类似于胚胎中的重要结构和过程 无疑在其他细胞的功能中扮演着重要的角色， 对这一模型系统的研究可能会揭示出 肌动蛋白的组织和功能将适用于所有细胞。 利用F-肌动蛋白在早期胚胎中鉴定的两种肌动蛋白相关蛋白 亲和层析并使用单抗在体内定位， 将进行详细研究。将任一种单抗注射到 早期胚胎会导致正常发育中断。它的作用机制 正在使用荧光标记的细胞骨架来研究破坏 蛋白质、组蛋白和抗体，使许多主要的 抗体作用期间的胚胎成分。这个 将使用标准生化技术从早期胚胎中提纯蛋白质 涉及肌动蛋白的体外活性的方法和特征 细丝。表达文库将使用抗体进行筛选 检测表达这些蛋白的cDNA克隆。一旦克隆编码 序列将被改变以确定蛋白质的哪些区域是 对功能很重要。将尝试进行体外和体内检测 以评估序列更改的影响。荧光标记和 注射改变的蛋白质将允许功能和定位 突变蛋白质将在体内进行检测。使用过度表达的蛋白质， 将构建亲和力矩阵以识别相互作用的组件 与这些蛋白质在被认为是大分子组装的 控制肌动蛋白在体内的聚合位置和时间。 对这些和其他控制细胞周期协调的蛋白质的研究 肌动蛋白在早期发育过程中的重排 果蝇胚胎可以为肌动蛋白的协调提供洞察 在细胞周期中与其他事件的重排和 肌动蛋白重排发生的机制。自从重新安排以来 在所有细胞中都是至关重要的，这些实验应该有助于 我们对肌动蛋白细丝结构在空间中定位的理解 并调节为在适当的时间聚合。

The early Drosophila embryo is being used to study the role of proteins that cooperate with actin to generate an organized cytoplasm in this large and highly dynamic syncytial cell. Actin filament structures participate in both the organization of the cytoplasm and the dynamic events required during normal development. Thus, these studies will provide insight into the actin-mediated process that contribute to the formation of a properly organized cellular blastoderm as a substrate of differentiation. Since actin structures and processes similar to those important in the embryo undoubtedly play important roles in the function of other cells, the studies of this model system will likely uncover general principles of actin organization and function that will apply to all cells. Two actin-associated proteins, identified in early embryos using F-actin affinity chromatography and located in vivo using monoclonal antibodies, will be studied in detail. Injections of either monoclonal antibody into early embryos causes disruption of normal development. The mechanism of disruption is being studied using fluorescently-labeled cytoskeletal proteins, histones and antibodies, allowing the visualization of many major embryonic components during the period when the antibody acts. The proteins will be purified from early embryos using standard biochemical methods and characterized for in vitro activities involving actin filaments. Expression libraries will be screened using the antibodies to detect cDNA clone that express these proteins. Once cloned coding sequences will be altered to determine which regions of the protein are important for function. Both in vitro and in vivo assays will be attempted to assess the impact of sequence changes. Fluorescent-labeling and injection of the altered protein will allow function and localization of mutant proteins to be assayed in vivo. Using overexpressed protein, affinity matricies will be constructed to identify componants that interact with these proteins in the macromolecular assemblies that are thought control actin polymerization sites and times in vivo. Study of these and other proteins that control the cell-cycle coordinated rearrangements of actin that occur during early development of the Drosophila embryo could provide insight into the coordination of actin rearrangements with the other events during the cell cycle and the mechanism by which actin rearrangements take place. Since rearrangements of actin are critical in all cells, these experiments should contribute to our understanding of how actin filament structures are positioned in space and regulated to polymerize at appropriate times.