Developmental function of Rb family proteins

描述（由申请人提供）：在控制细胞增殖和分化的许多途径中，视网膜母细胞瘤蛋白（Rb）调控网络的基因在肿瘤发生期间作为突变或失调的频繁靶点（如果不是强制性靶点）而突出。虽然生物化学和组织培养的研究涉及Rb家族成员在广泛的细胞活动，真正的功能Rb在体内和正常发育过程中没有得到很好的理解。我们的长期目标是在分子水平上了解Rb家族蛋白的细胞和发育功能。为此，我们设计了一种遗传策略，使我们能够识别与C。elegans Rb同源物lin-35，控制重要的发育过程。使用这个系统，我们已经证明了典型的细胞周期功能的LIN-35，以及一个新的作用，这种蛋白质在器官形态发生。我们还发现了一个互补的途径，通过UBC-18/UbcH 7控制器官形态发生，UBC-18/UbcH 7是一种保守的泛素缀合酶，参与靶向蛋白质降解。这些实验旨在揭示LIN-35与一个或多个平行通路共同作用调节C.优雅 我们的主要目标分为两类。一个广泛的目标是确定与LIN-35和UBC-18协同作用以控制器官发生的其他因子。这些研究将包括sir-9的克隆和表征，sir-9是一种基因，与ubc-18一样，与lin-35冗余地起作用以控制器官形态发生;执行双杂交筛选以鉴定UBC-18相互作用蛋白;以及使用已知或推定的泛素途径组分进行定向RNAi喂养筛选。我们的第二个目标是确定LIN-35和UBC-18调控的功能相关下游靶标。这些研究将包括遗传选择，以分离抑制lin-35致死性的突变; ubc-18双突变体;微阵列分析，以确定LIN-35调节的转录物的完整谱;以及使用通过早期双杂交或RNAi喂养筛选确定的UBC-18辅助因子的额外双杂交筛选。这些研究的成功完成将大大提高我们对Rb家族功能的总体了解，并将为Rb蛋白在形态发生中的新作用提供详细的机制知识。

DESCRIPTION (provided by applicant): Among the many pathways controlling cell proliferation and differentiation, genes of the retinoblastoma protein (Rb) regulatory network stand out as frequent if not obligatory targets for mutation or deregulation during tumorigenesis. Although biochemical and tissue culture studies have implicated Rb family members in a wide range of cellular activities, the bona fide functions of Rb in vivo and during normal development are not well understood. Our long-term objective is to understand the cellular, and developmental functions of Rb family proteins at the molecular level. To this end, we have devised a genetic strategy that has allowed us to identify genes and pathways that function coordinately with the C. elegans Rb homolog, lin-35, to control essential developmental processes. Using this system, we have demonstrated canonical cell cycle functions for LIN-35 as well as a novel role for this protein in organ morphogenesis. We have also uncovered a complementary pathway that acts to control organ morphogenesis through UBC-18/UbcH7, a conserved ubiquitin-conjugating enzyme involved in the targeting of proteins for degradation. The proposed experiments are designed to uncover the underlying mechanism by which LIN-35, acting in conjunction with one or more parallel pathways, regulates organ morphogenesis in C. elegans. Our main objectives fall into two categories. One broad aim is to identify additional factors that function cooperatively with LIN-35 and UBC-18 to control organogenesis. These studies will include the cloning and characterization of sir-9, a gene that, like ubc-18, functions redundantly with lin-35 to control organ morphogenesis; the execution of a two-hybrid screen to identify UBC-18-interacting proteins; and a directed RNAi feeding screen using known or putative ubiquitin pathway components. Our second objective is to identify functionally relevant downstream targets for regulation by LIN-35 and UBC-18. These studies will include genetic selections to isolate mutations that suppress the lethality of lin-35; ubc-18 double mutants; microarray analyses to identify the complete spectrum of LIN-35-regulated transcripts; and additional two-hybrid screens using co-factors of UBC-18 identified through earlier two-hybrid or RNAi-feeding screens. The successful completion of these studies will greatly enhance our general understanding of Rb family functions and will provide detailed mechanistic knowledge of this novel role for Rb proteins in morphogenesis.