Activation of Phospholipase Cbeta by G Proteins

描述（由申请人提供）：P.I. S. Scarlata磷脂酶C-b（PLC？）酶被G蛋白激活以响应诸如激素和神经递质的试剂。PLC？活性引起细胞内钙的增加，这最终导致细胞的深刻变化。我们的实验室一直致力于研究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，膜域称为小窝可以控制PLC的持续时间？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，以确定通过哪些机制小窝caveolae PLC？- 介导的钙信号并沿沿着特定的细胞途径引导信号。2-我们已经发现PLC b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。translin相关蛋白X（TRAX）。TRAX及其伴侣translin是调节蛋白质细胞水平的siRNA机制的主要组分。在AIM 2中，我们将确定TRAX提供PLC的能力？从细胞核转移到质膜。同时，我们将确定PLC是否影响translin-TRAX功能。3-PLC的能力？产生信号取决于它的细胞浓度。我们已经发现突触核蛋白可以结合并稳定PLC？在细胞中。α-突触核蛋白（AS）是一种功能未知的未折叠小蛋白，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，下调PLC？？导致AS聚集。此外，γ-突触核蛋白（GS），乳腺癌易感基因1编码的蛋白质，似乎增加PLCb介导的迁移和细胞侵袭，从而促进乳腺癌。在AIM 3中，我们将更好地定义PLC？- 突触核蛋白相互作用和开发防止AS聚集和减少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.