Activation of Phospholipase Cbeta by G Proteins

描述(申请人提供)：P.I.S.斯卡拉塔磷脂酶C-b(PLC？)酶被G蛋白激活，以响应激素和神经递质等因素。PLC？活动导致细胞内钙的增加，最终导致细胞的深刻变化。本实验室主要研究G蛋白激活PLC的机制。在分子水平上。然而，在培养的细胞中，我们发现了额外的和意想不到的调节机制。首先，被称为小凹的膜域能控制PLC的持续时间吗？通过选择性地隔离G蛋白激活剂来激活。1-在AIM 1中，我们将使用活细胞成像和最先进的荧光方法来确定小凹延长PLC？介导的钙信号并沿着特定的细胞路径引导信号的机制。2-我们发现PLCB定位于细胞核和胞浆以及质膜，在那里发现了它的G蛋白激活剂。我们发现了一种新的PLC蛋白伴侣？转化素相关蛋白X(Trax)。Trax及其伙伴翻译蛋白是调节细胞蛋白质水平的siRNA机制的主要组成部分。在AIM 2中，我们将确定Trax交付PLC的能力？G蛋白被激活后，从细胞核到质膜。同时，我们将确定PLC是否会影响Translin-Trax功能。3-PLC的能力？产生信号取决于它的细胞浓度。我们发现突触核蛋白可以与PLC结合并稳定下来。在牢房里。α-突触核蛋白(AS)是一种功能未知的小分子未折叠蛋白，是神经退行性斑块的主要成分。AS的存在大大增加了PLC？水平，我们发现PLC？？的下调是由于聚集。此外，乳腺癌易感基因1编码的蛋白-乳腺癌易感基因1编码的蛋白质在目标3中，我们将更好地定义PLC？-突触核蛋白的相互作用，并开发防止AS聚集和降低GS诱导的乳腺癌表型的试剂。

DESCRIPTION (provided by applicant): P.I. S. Scarlata Phospholipase C-b (PLC?) enzymes are activated by G proteins in response to agents such as hormones and neurotransmitters. PLC? activity causes an increase in intracellular calcium which ultimately leads to profound changes in the cell. Our lab has focused on the mechanism through which G proteins activate PLC? on the molecular level. However, in cultured cells, we find additional and unexpected mechanisms of regulation. First, membrane domains called caveolae can control the duration of PLC? activation by selectively sequestering G protein activators. 1 - In AIM 1, we will use live cell imaging and state of the art fluorescence methods to determine the mechanisms through which caveolae prolongs PLC? - mediated calcium signals and directs signals along specific cellular pathways. 2- We have found that PLCb localizes to the nucleus and cytosol as well as the plasma membrane where its G protein activators are found. We have identified a novel protein partner of PLC? translin-associated protein X (TRAX). TRAX and its partner translin are major components of the siRNA machinery regulating the cellular levels of proteins. In AIM 2, we will determine the ability of TRAX to deliver PLC? from the nucleus to the plasma membrane upon G protein activation. In parallel, we will determine whether PLC affects translin-TRAX function. 3- The ability of PLC? to generate signals depends on its cellular concentration. We have found that the synucleins can binds to and stabilize PLC? in cells. Alpha-synuclein (AS) is a small unfolded protein of unknown function and is a major component of neurodegenerative plaques. The presence of AS greatly increases PLC? levels, and we find that down-regulation of PLC??causes AS aggregation. Also, gamma- synuclein (GS), the protein encoded by the breast cancer susceptibility gene 1, appears to increase PLCb -mediated migration and cell invasiveness thereby promoting breast cancer. In AIM 3 we will better define PLC? - synuclein interactions and develop reagents that prevent AS aggregation and reduce GS-induced breast cancer phenotype.