Cytoplasmic Retention as a Regulatory Mechanism in Embryogenesis

9319178 Etkin许多核蛋白的功能受选择性核定位的调节。非洲爪哇核因子7(Xnf7)是一个新的锌指基因家族成员，其成员主要由转录因子和原癌基因组成。XNF7首先在卵母细胞核中检测到，并保留在细胞质中，直到发育中期的囊胚重新进入细胞核。最近的证据表明，XNF7中一个称为细胞质保留结构域(CRD)的22个氨基酸序列是囊胚期中期细胞质保持所必需的，并且蛋白质中两个位点的磷酸化是CRD正常功能所必需的。埃特金博士的实验室表明，XNF7通过一种锚定机制保留在细胞质中，这种锚定机制很可能涉及到与胞浆锚定蛋白的连接。本项目检验的假设是，磷酸化诱导XNF7的结构变化，导致CRD序列与锚蛋白结合。埃特金博士将通过实现以下具体目标来检验这一假说：(1)他将进一步确定磷酸化在细胞质中保留xnf7的作用。(2)他将分析和鉴定与CRD序列相互作用并在细胞质中保留XNF7功能的蛋白质，并克隆这些蛋白质的cDNA。除了深入了解两栖动物发育过程中的一个重要调控过程外，对XNF7选择性核定位机制的分析将为理解其他系统中的这一现象提供必要的信息。***

9319178 Etkin The function of many nuclear proteins is regulated by selective nuclear localization. Xenopus nuclear factor 7 (xnf7), a putative transcription factor, is a member of a novel zinc finger gene family whose members consist primarily of transcription factors and protooncogenes. Xnf7 is first detected in the oocyte nucleus and is retained in the cytoplasm until the mid blastula stage of development when it reenters the nucleus. Recent evidence showed that a 22 amino acid sequence in xnf7 called the cytoplasmic retention domain (CRD) is required for cytoplasmic retention prior to the mid blastula stage and that phosphorylation of two sites within the protein is necessary for proper CRD function. Dr. Etkin's laboratory has shown that xnf7 is retained in the cytoplasm through an anchor mechanism most likely involving the tethering to a ctyoplasmic anchor protein. The hypothesis being tested in this project is that phosphorylation induces a structural change in xnf7 resulting in the binding of the CRD sequence to an anchor protein. Dr. Etkin will test this hypothesis by accomplishing the following specific aims: (1) He will further determine the role of phosphorylation in the retention of xnf7 in the cytoplasm. (2) He will analyze and identify the proteins that interact with the CRD sequence and function in cytoplasmic retention of xnf7 and will clone the cDNAs for these proteins. In addition to gaining an insight into an important regulatory process during amphibian development, the analysis of the mechanism of selective nuclear localization of xnf7 will provide information necessary to understand this phenomenon in other systems. ***