Alternative mechanisms of signaling via trimeric G proteins

描述(由申请人提供)：在这里，我们建议鉴定和描述一类对细胞通讯的核心机制至关重要的新的信号分子，即通过三聚体G蛋白的信号。三聚体G蛋白在信号转导中起着关键作用 这种机制从酵母到人类都是保守的，构成了25%上市药物的靶标。尽管在过去的几十年里取得了重大进展，但对这一信号机制的理解仍然不完整--控制三聚体G蛋白的调控网络已经超出了这一信号通路的传统观点所提出的范围。这表明，三聚体G蛋白的调节比以前所认识的要复杂得多，新的治疗靶点可能来自G蛋白调节的替代机制的发现。我们的目标是通过识别一个新的激活剂家族，剖析它们与G蛋白偶联的分子基础，并表征该家族的原型成员促进癌症进展向转移的基本分子机制，来进一步了解G蛋白调控网络。根据我们和其他人以前的观察，我们假设一个新的非受体G蛋白激活剂家族是由鸟嘌呤核苷酸交换因子(Global)基序的存在定义的，这些激活剂组装与疾病相关的替代G蛋白信号通路。这些新的激活物不一定是膜蛋白，可以替代或平行于典型的激活物，即G蛋白偶联受体(GPCRs)，从而代表了对这一信号通路的经典观点的迂回。SA#1中的实验旨在通过生物信息学、高通量多肽阵列筛选和酵母功能分析来鉴定和表征新型非受体G蛋白激活剂。我们已经通过识别和部分描述一些预测的候选对象的生理功能来验证这一总体方法。在SA#2中，我们建议进行实验，以了解这一新的非受体GEF家族如何结合和激活G蛋白的结构基础。综上所述，这些研究将确定定义一个全新的G蛋白调节剂家族的基本分子原理，并将产生未来对它们进行机制研究所需的工具。SA#3中提出的实验测试了一般假设，即这些非受体GEF在疾病中组装了针对GIV特定病例的替代信号电路，GIV是这一新的GEF家族的原型成员。到目前为止，我们的工作提出了一个模型，在该模型中，需要依赖GIV激活G蛋白来组装一条信号通路，将细胞外基质蛋白对整合素的刺激与肿瘤细胞获得促转移特征联系起来。综上所述，我们希望通过结合基于发现的和有针对性的机制研究来揭开G蛋白调节的新的一般机制，在疾病中具有广泛的意义。

DESCRIPTION (provided by applicant): Here we propose to identify and characterize a new class of signaling molecules critical for a core mechanism of cellular communication, i.e., signaling via trimeric G proteins. Trimeric G proteins play a pivotal role in a signal transduction mechanism that is conserved from yeast to humans and that constitutes the target for >25% of marketed drugs. Despite major advances in the last decades, the understanding of this signaling mechanism remains incomplete- the network of regulators that control trimeric G proteins has expanded beyond what the traditional view of this signaling pathway proposes. This has made evident that the regulation of trimeric G proteins is more complex than previously appreciated and that novel therapeutic targets may arise from the discovery of alternative mechanisms of G protein regulation. Our goal is to further the understanding of the G protein regulatory network by identifying a new family of activators, dissecting the molecular basis of their coupling to G proteins and characterizing the basic molecular mechanisms by which the prototype member of this family promotes cancer progression towards metastasis. Based on previous observations by us and others we hypothesize that a new family of non-receptor G protein activators is defined by the presence of a guanine nucleotide exchange factor (GEF) motif and that these activators assemble alternative G protein signaling circuits involved in disease. These new activators are not necessarily membrane proteins and may work in lieu of or in parallel to the canonical activators, i.e., G protein-coupled receptors (GPCRs), thereby representing a detour from the classical view of this signaling pathway. The experiments in SA#1 are designed to identify and characterize novel non-receptor G protein activators with a GEF motif by using bioinformatics, high-throughput peptide array screening and functional assays in yeast. We have validated this overall approach by identifying and partially characterizing the physiological function for some of the predicted candidates. In SA#2 we propose experiments to understand the structural basis for how this new family of non-receptor GEFs bind and activate G proteins. Taken together, these studies will determine the basic molecular principles that define a whole new family of G protein regulators and will also generate the tools required for future mechanistic studies on them. Experiments proposed in SA#3 tests the general hypothesis that these non-receptor GEFs assemble alternative signaling circuits in disease for the particular case of GIV, the prototype member of this new family of GEFs. Our work to date suggests a model in which GIV-dependent activation of G proteins is required to assemble a signaling pathway linking stimulation of integrins by extracellular matrix proteins to the acquisition of pro-metastatic features by tumor cells. In summary, we hope to unravel a new general mechanism of G protein regulation with broad implications in disease by combining discovery-based and targeted mechanistic studies.