CONTROLLING THE ACTIVITY OF A DROSOPHILA TGF-B HOMOLOG

迈克尔B博士。奥康纳目前是一个助理教授在 分子生物学和生物化学系 加州-欧文。 候选人的长期研究兴趣是 建立如何在分子水平上指定位置信息 并在开发过程中进行解释。 果蝇Drosophila Melanogaster 将被用作实验系统。 当前的目标是 研究tolloid（tld）、螺钉（scw）和tld所起的作用 相关的（tld-r）基因产物（tlc，SCW，TLD-R）在指导 背/腹（D/V）模式的形成在发育中的胚胎。 遗传 分析表明，TLD和SCW都能促进 第三个D/V模式基因decapentaplegic（dpp），其产物（DPP）是一个 生长因子的TGF-β超家族成员。 tgf-β家族 的分泌多肽介导细胞间通讯， 许多或生物体，似乎控制了广泛的生物 包括细胞生长和分化的过程。 为了澄清 的tld-dpp和scw-dpp相互作用，结构，表达， 将检测这些基因产物的生物化学活性。 对于TLD A 杆状病毒表达系统将用于过表达蛋白质。 DNA序列分析显示，tld与人类骨骼有45%的相同性 形态发生蛋白-1（BMP-1），一种假定的金属蛋白酶。 纯化的葡萄糖 将使用放射性标记的酪蛋白检测蛋白质的蛋白水解活性 和香豆素衍生肽作为底物。 搜索TLD-DPP 复合物，抗体将针对不同的部分， 用于免疫共沉淀实验。 切割的能力 将通过共表达蛋白的SDS-PAGE电泳监测DPP。 为了将特定的tld结构域与功能联系起来， 有11种不同的tld突变，这些突变会导致与 dpp，将通过DNA测序鉴定。 这项工作需要 博士奥康纳获得了一些新的实验技能，包括 纯化蛋白质，制造抗体，并开发生物化学分析， 蛋白质功能 这个部门有几个资深的调查员 包括Krishna Tewari博士，Agnes Henschen博士，Charles Glabe博士和Dr. 芭芭拉伯吉斯，她的研究利用各种技术在蛋白质 每天都有化学反应 在他们的指导下，奥康纳医生 发展必要的技能，以胜任解决蛋白质问题 结构和功能。 从长远来看，充分利用 基因、分子和生物化学技术对实验的补充 这些问题将大大提高奥康纳博士的研究能力。 系里将解除候选人的教学和委员会职务 在获奖期间，他可以把他的全部责任， 努力使自己成为一名独立调查员。

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.