CONTROLLING THE ACTIVITY OF A DROSOPHILA TGF-B HOMOLOG

迈克尔·B·奥康纳博士目前是 加州大学分子生物学与生物化学系 加州--欧文。候选人的长期研究兴趣是 在分子水平上确定位置信息是如何指定的 并在开发过程中进行解释。果蝇--黑腹果蝇 将被用作实验系统。眼前的目标是 调查Tolloid(TLD)、螺钉(SCW)和TLD所起的作用 相关(TLD-r)基因产物(TLD、SCW、TLD-R) 发育中胚胎的背侧/腹侧(D/V)模式的形成。遗传 分析表明，TLD和SCW都能促进 第三种D/V模式基因DPP，其产物(DPP)是 生长因子超家族中的一员。转化生长因子-β家族 分泌多肽在细胞间通讯中的作用 大量或有机体，似乎控制着广泛的生物 包括细胞生长和分化的过程。为了澄清本质 TLD-DPP和SCW-DPP相互作用的结构、表达和 将对这些基因产物的生化活性进行检测。对于TLD a 将利用杆状病毒表达系统过表达该蛋白。 DNA序列分析表明，TLD与人骨的同源性为45% 形态发生蛋白-1(BMP-1)，一种可能的金属蛋白酶。提纯热释光二极管 将使用放射性标记的酪蛋白来测试蛋白质的分解活性 和香豆素衍生肽作为底物。搜索TLD-DPP 复合体，抗体将针对TLD的不同部分和 用于免疫共沉淀实验。TLD的切割能力 DPP将通过共表达蛋白的SDS-PAGE电泳法进行监测。 为了将特定的TLD结构域与功能相关联，皮损与 有11种不同的TLD突变，导致与 DPP，将通过DNA测序进行鉴定。这项工作将使 奥康纳博士获得了几项新的实验技能，包括 提纯蛋白质，制造抗体，并开发生化分析方法 蛋白质的功能。该部门有几名资深调查人员 包括Krishna Tewari博士、Agnes Henschen博士、Charles Glabe博士和Dr. Barbara Burgess，她的研究利用了蛋白质的各种技术 每天的化学反应。在他们的指导下，奥康纳博士将 发展必要的技能，以熟练地解决蛋白质中的问题 结构和功能。从长远来看，完全应用 遗传学、分子和生化技术对实验的补充 这些问题将大大增强奥康纳博士的研究能力。 该系将免除候选人的教职和委员会职务。 在颁奖期间的责任，以便他可以全力以赴 努力使自己成为一名独立的调查员。

Dr. Michael B. O'Connor is currently an assistant professor in the Department of Molecular Biology and Biochemistry at the University of California - Irvine. The candidates long term research interest is to establish how, at the molecular level, positional information is specified and interpreted during development. The fruit fly Drosophila Melanogaster is to be used as the experimental system. The immediate aim is to investigate the roles played by the tolloid (tld), screw (scw), and tld related (tld-r) gene products (TLD, SCW, TLD-R) in the process that directs dorsal/ventral (D/V) pattern formation in the developing embryo. Genetic analysis suggests that both tld and scw act to boost the activity of a third D/V patterning gene decapentaplegic (dpp), whose product (DPP) is a member of the TGF-beta superfamily of growth factors. The TGF-beta family of secretory polypeptides mediate intercellular communication in a multitude or organisms and appear to control a wide range of biological processes including cell growth and differentiation. To clarify the nature of tld-dpp and scw-dpp interactions, the structure, expression, and biochemical activities of these gene products will be examined. For tld a baculovirus expression system will be used to over-express the protein. DNA sequence analysis has revealed that tld is 45% identical to human bone morphogenetic protein-1 (BMP-1), a putative metalloprotease. Purified TLD protein will be tested for proteolytic activity using radiolabeled casein and coumarin derivatized peptides as substrates. To search for TLD-DPP complexes, antibodies will be raised against different parts of TLD and used in co-immunoprecipitation experiments. The ability of TLD to cleave DPP will be monitored by SDS-PAGE electrophoresis of co-expressed proteins. To correlate specific tld domains with function, the lesions associated with 11 different tld mutations, which cause abnormal interactions with dpp, will be identified by DNA sequencing. this work will necessitate that Dr. O'Connor acquire several new experimental skills including the ability to purify proteins, make antibodies, and develop biochemical assays for protein function. The department has several established investigators including Dr. Krishna Tewari, Dr. Agnes Henschen, Dr. Charles Glabe and Dr. Barbara Burgess, whose research utilizes various techniques in protein chemistry on a daily basis. Under their tutelage, Dr. O'Connor will develop the necessary skills to competently address problems in protein structure and function. In the long term, the ability to apply a full complement of genetic, molecular and biochemical techniques to experimental problems will significantly enhance Dr. O'Connor's research capabilities. The department will relieve the candidate from teaching and committee responsibilities during the award period so that he can devote his full efforts towards establishing himself as an independent investigator.