GS PATHWAYS IN DROSOPHILA EPITHELIAL CELLS

描述：本提案的目标是定义如何激活 果蝇G蛋白Gsα亚单位(DGs)对 利用翼泡作为一种特殊细胞类型的发育和功能 范例。研究人员使用一种复杂的体内表达系统 基于GAL4控件元素，已注意到 Dgs的激活突变(Dgs*)会导致众多复杂的表型。 最简单也是最稳定的观察之一就是机翼的形成 介导翼上皮细胞表达的GAL4系引起的水泡 幼虫期晚期。据推测，起泡是由于断裂引起的 整合素介导的粘连失败所致的经翼连接 在机翼扩张过程中。非常有趣的是，依赖DGs*的 即使在没有蛋白激酶A(零突变)的情况下也会发生水泡， 提示DGs*使用cAMP非依赖途径或cAMP，但 而不是PKA。第一个目标是检验这样一种假设，即营地的上升 翅上皮细胞是水泡产生所必需的。一个 一种基于A基因表达的细胞内cAMP水平监测系统 将建立β-半乳糖苷酶报告，然后对cAMP进行调节 通过过表达腺酰环化酶(Rutabaga)或 磷酸二酯酶(DUNCE)。第二个目标将是识别分子 它们是翼上皮细胞中DGs*激活途径的组成部分。 研究人员假设，调节下游的分子子集 影响存在于关键水平，并将利用以下事实 果蝇的大多数基因在发育缺陷时是剂量补偿的 筛选识别当以一个副本存在时可以抑制的基因 水泡的形成。第三个目标是测试这样一种想法，即 翼上皮细胞内的Gs途径是调节粘附性的途径 整合素分子的性质。翅膀形态发生的特定方面 将会被检验，基因操作的能力也会被检验 编码整合素蛋白以促进或抑制水泡的形成。

DESCRIPTION: The objective of this proposal is to define how the activation of the G protein Gs alpha subunit (DGs) in Drosophila impacts on the development and function of a specific cell type using wing blistering as a paradigm. The investigator, using a sophisticated in vivo expression system based on GAL4 control elements, has noted that the expression of an activated mutant of DGs (DGs*) results in numerous and complex phenotypes. One of the simplest and most consistently observed is the formation of wing blisters by GAL4 lines that mediate expression in wing epithelium during late pupal periods. The blistering is proposed to occur from rupture of transalar connections through the failure of integrin-mediated adhesion processes during wing expansion. Of great interest, the DGs*-dependent blistering occurs even in the absence (null mutation) of protein kinase A, suggesting that DGs* utilizes either a cAMP-independent pathway or cAMP but not PKA. The first aim is to test the hypothesis that elevation of cAMP in wing epithelial cells is required for the generation of blistering. A system for monitoring intracellular cAMP levels based on expression of a beta-galactosidase reporter will be established, then cAMP will be modulated through overexpression of an adenylyl cyclase (rutabaga) or phosphodiesterase (dunce). The second aim will be to identify molecules which are components of the DGs*-activated pathway in wing epithelial cells. The investigator assumes that a subset of molecules mediating downstream effects exist at a critical level and will take advantage of the fact that most genes in Drosophila are dosage-compensated, in developing deficiency screens to identify genes which, when present in one copy, can suppress blister formation. The third aim is to test the idea that the function of the Gs pathway within wing epithelial cells is to regulate the adhesive properties of integrin molecules. Specific aspects of wing morphogenesis will be examined, as will the ability of genetic manipulations of genes encoding integrin proteins to enhance or suppress blister formation.