IMMUNOLOCALIZATION OF G PROTEINS IN MAMMALIAN CELLS

异源三聚体GTP结合蛋白（G蛋白）在 在跨质膜传递受体产生的信号方面。 其中一个最好的特征信号转导途径， 腺苷酸敏感环化酶系统。 效应酶在 双重刺激和抑制控制的两个G蛋白称为G和 G. 然而，现在已知这些G蛋白中的每一种都在以下方面发挥作用： 除了调节腺苷酸环化酶，在G的情况下， 是三个密切相关的基因产物，可以在一个单一的表达， 细胞的类型 G的每种不同形式的函数都不是 知道的 对信号转导机制的理解是基础 包括激素和药物作用在内的医学的许多方面。 改变 G蛋白的活性已经在诸如心力衰竭， 高血压和癌症。 了解特定G的位置 蛋白质可以提供对这种疾病的临床相关性的了解， 观察. 关于G的亚细胞分布的大部分信息 蛋白质来自于相对粗糙的尝试， 分馏，并有很少的超微结构的信息，他们的 重要课题。 G蛋白活性，虽然明显存在于 据报道，质膜与细胞内 隔室，包括高尔基体，线粒体，内体， 细胞核，激活后，细胞质。1)第一特定 目的是利用免疫电子显微镜的冷冻切片， 定量鉴定G. 蛋白质亚基在细胞中是相关的。 将试图 确定质膜上的专门结构位点， 由G和G的每种形式填充的细胞内膜。 的 待检验假设是个体的帽子差异分布 G蛋白寡聚体有助于其参与的特异性 在信号转导途径和G蛋白知识方面 本地化将有助于了解他们的职能作用。2)第二 具体目标包括审查界定的结构性 α亚基在膜定位中的特征。 分布 内源性G蛋白将使用定量 全细胞和血浆的形态学和生化技术 膜暴露于已知改变G蛋白功能的试剂。 总部设在 在这些实验的结果中，突变的G蛋白α亚基 处于活动和非活动状态。 3)第三个具体目标是 确定受体定位与 它们的附属G蛋白 将检查受体和G蛋白 对于基底侧或顶端质膜的差异分布 的上皮细胞。 此外，G在脱敏过程中的命运 可以提供洞察G蛋白的其他功能，以及 解释G蛋白在信号传导中的特异性的机制。

Heterotrimeric GTP binding proteins (G proteins) serve an essential role in transducing receptor-generated signals across the plasma membrane. One of the best characterized signal transduction pathways in the hormone-sensitive adenylyl cyclase system. The effector enzyme is under dual stimulatory and inhibitory control by two G proteins termed G and G. However, it is now known that each of these G proteins has roles in addition to regulation of adenylyl cyclase and, in the case of G there are three closely related gene products that can be expressed in a single type of cell. The function of each of the different forms of G is not known. An understanding of signal transduction mechanisms is fundamental to many aspect of medicine including hormone and drug action. Altered G protein activity has been observed in diseases such as heart failure, hypertension, and cancer. Knowledge of the location of specific G proteins may provide insight into the clinical relevance of such observation. Most information about the subcellular distribution of G proteins has come from relatively crude attempts at cellular fractionation, and there is little ultrastructural information on their important subject. G protein activity, although clearly present in the plasma membrane, has been reported to be associated with intracellular compartment, including the Golgiapparatus,mitochondria, endosomes, nucleus, and following activation, the cytoplasm. 1) The first specific aim is to utilize immunoelectron microscopy of ultrathin frozen sections to identify quantitatively the subcellular structures with which G protein subunits are associated in cells. Attempts will be made to identify specialized structural sites on the plasma membrane and intracellular membranes that are populated by G and each form of G. The hypothesis to be tested is hat differential distribution of individual G protein oligomers contributes to the specificity of their participation in signal transduction pathways and that knowledge of G protein localization will shed light on their functional roles. 2) The second specific aim involves examination of the role of defined structural features of alpha subunits in membrane localization. The distribution of endogenous G proteins will be examined using quantitative morphological and biochemical techniques on whole cells and plasma membranes exposed to agents known to alter G protein function. Based in part on the results of these experiments, mutant G protein alpha subunits in the active and inactive states. 3) The third specific aim is to determine the relationship between the localization of receptors and their affiliated G proteins. Receptors and G proteins will be examined for differential distribution to basolateral or apical plasma membranes of epithelial cells. In addition, the fate of G during desensitization may provide insight into additional functions of G proteins as well as mechanisms to explain G protein specificity in signal transduction.