KINASES AND CONTROL OF CELL-TYPE SPECIALIZATION

已知蛋白激酶在调节中起关键作用 级联对控制动物细胞增殖和 专业化。这样的监管网络的医学相关性 最值得欣赏的是考虑到转换后的状态 细胞失去了部分分化的表型， 扩散到外部或正常的监管限制。蛋白 激酶是已被确认为 负责转型的协作代理与 各种癌症。 酿酒酵母(S.cerevisiae)是研究酿酒酵母的模式系统。 控制细胞类型特化的分子机制。我们的 目的是确定两种蛋白激酶STE7的作用 以及Stell基因产品，在细胞特化方面。我们会 调查：(1)影响时序和/或细胞类型的因素 蛋白激酶活性的特异性(2)结构变化 在蛋白激酶中，可以改变它们的控制、特异性和 功能和(3)控制电路中相互作用的组件 蛋白激酶作为底物或调节剂。《原则》 此处未涵盖的内容应适用于在 多细胞真核生物。 为此目的，将STE基因产物在大肠杆菌中表达 对酶的活性和酶学性质进行了纯化和评价。 大肠杆菌表达的材料也将被用来诱导 兔抗血清。STE特异性抗体将被用作 检测STE基因产物细胞类型特异性的试剂 活动、修改或本地化。体外诱变将 应用于在STE基因中产生结构改变 可能会阻止功能或以其他方式改变特异性的产品 在体内发挥作用。利用In进行伪回归研究 体外产生的STE突变等位基因将用于鉴定 交配调控途径的其他组成部分。

Protein kinases are known to play a pivotal role in regulatory cascades important to the control of animal cell proliferation and specialization. The medical relevance of such regulatory networks is best appreciated by considering the transformed state where cells have lost aspects of their differentiated phenotype and proliferate outside or normal regulatory constraints. Protein kinases are_among the "oncogenes" that have been identified as collaborative agents responsible for transformation associated with various cancers. The yeast, S. cerevisiae is a model system for the study of molecular mechanisms controlling cell-type specialization. Our objective is to define the role of two protein kinases, the STE7 and STEll gene products, in cell specialization. We will investigate: (1) factors that modulate the timing and/or cell-type specificity of protein kinase activity (2) structural alterations in protein kinases that can modify their control, specificity and function and (3) components in control circuits that interact with protein kinases as substrates or as regulators. The principles uncovered here should be applicable to pathways operating in multicellular eukaryotes. The STE gene products will be expressed in E. coli for the purpose of purification and evaluation of kinase activity and enzymology. The E. coli expressed material will also be used to elicit antisera in rabbits. The STE specific antibodies will be used as reagents to determine cell-type specificity of STE gene product activity, modification or localization. In vitro mutagenesis will be applied to create structural alterations in the STE gene products that may prevent function or otherwise modify specificity of function in vivo. Pseudoreversion studies utilizing the in vitro generated STE mutant alleles will be applied to identify other components of the mating regulatory pathway.