RETINOBLASTOMA PROTEIN IN DROSOPHILA

该研究的长期目标是阐明控制细胞增殖和分化的机制。具体来说，本基金的研究调查了视网膜母细胞瘤（Retinoblastoma， RB）蛋白家族在发育过程中的功能和调控。RB家族被认为在细胞增殖和细胞分化中发挥重要作用。了解这一特殊蛋白质家族的正常功能将增强我们对细胞增殖和分化是如何被控制的总体理解。拟开展的研究主要集中在果蝇遗传系统中的RB家族同源。尽管RB-E2F途径已经通过生化方法进行了广泛的研究，但RB、E2F和DP家族蛋白质的体内功能及其功能相互作用的研究受到每个家族中大量蛋白质的存在和缺乏有效的方法来功能鉴定该途径中的基因的阻碍。果蝇RB途径的研究规避了这些问题，并补充了标准的生化方法。果蝇RB通路的更简单的图像，以及在此类研究中易于结合遗传，分子，细胞和发育方法，使果蝇成为开展所提出的研究的理想模型系统。在初步研究中，我们克隆了一个果蝇RB家族同源RBF，并证明了RB- e2f通路的功能和调控在果蝇和哺乳动物之间是保守的。此外，我们已经为拟议的研究生成了许多试剂，包括：RBF突变体，抗RBF单克隆抗体和一些RBF过表达菌株。此外，还建立了基因筛选来鉴定RB通路中的基因。在这项资助中，我们建议使用这个果蝇遗传系统来研究RBF的体内功能，并获得RB途径的功能解剖。具体而言，我们提出(i)研究RBF在发育过程中的功能，（ii）通过挽救RBF突变体的致死性来分析RBF的结构和功能，（iii）研究E2F与RBF、Dacapo与RBF之间的功能关系，（iv）继续进行RB通路基因筛选。

The long-term objective of the proposed research is to elucidate the mechanisms by which cell proliferation and differentiation are controlled. Specifically, the research in this grant investigates the function and regulation of the Retinoblastoma (RB) family of proteins during development. The RB family is thought to play important roles in both cell proliferation and in cell differentiation. Understanding the normal function of this particular family of proteins will enhance our general understanding of how cell proliferation and differentiation are controlled. The proposed studies focus on the RB family homolog in a Drosophila genetic system. Although the RB-E2F pathway has been extensively studied using biochemical approaches, studies of the in vivo function of RB, E2F, and DP family of proteins and their functional interactions have been hampered by the presence of large number of proteins in each family and the lack of efficient approaches to functionally identify genes in the pathway. Studies of the RB pathway in Drosophila circumvents these problems and complements standard biochemical approaches. The much simpler picture of the RB pathway in Drosophila and the ease of combining genetic, molecular, cellular, and developmental approaches in such studies make Drosophila an ideal model system to carry out the proposed research. In preliminary studies we have cloned a Drosophila RB family homolog RBF and have demonstrated that the function and regulation of the RB-E2F pathway is conserved between Drosophila and mammals. Furthermore we have generated a number of reagents for the proposed research including: RBF mutants, monoclonal antibodies against RBF, and a number of RBF overexpression strains. In addition, genetic screens have been set up to identify genes in the RB pathway. In this grant we propose to use this Drosophila genetic system to study the in vivo function of RBF and to obtain a functional dissection of the RB pathway. Specifically, we propose (i) to study the function of RBF during development, (ii) to analyze the structure and function of RBF through rescuing the lethality of RBF mutant, (iii) to investigate the functional relationship between E2F and RBF, and between Dacapo and RBF, (iv) to continue our genetic screen to identify genes of the RB pathway.