Molecular Mechanisms of Cell Invasion

描述（由申请人提供）：这是一项研究侵袭性肿瘤细胞中微泡生物发生机制的应用。它建立在先前NIH资助的一个独特的囊泡群体上产生的令人兴奋的发现的基础上，这些囊泡被称为微囊泡，含有功能活性蛋白酶，并在肿瘤细胞获得侵袭潜力时被释放。负载蛋白酶的微泡的释放可能作为一种机制，导致基质降解，甚至可能在远端位置沉积存款旁分泌信息，从而在肿瘤细胞入侵并迁移通过周围组织时创建“最小阻力”的路径。这与侵入伪足处的细胞周蛋白水解不同，侵入伪足处的细胞周蛋白水解使得与前缘并列的局部基质降解成为可能。发现可能存在一种以上的蛋白水解侵袭模式，限制了不包括局灶性和远端蛋白水解的任何侵袭靶向治疗策略的有效性。虽然大量的研究已被导向了解侵袭伪足的形成机制和在细胞侵袭位点的功能，微泡的生物发生和功能仍然是肿瘤生物学的一个相对不足的领域。然而，最近越来越多的证据表明，真正存在的微泡在体液（血液，尿液和腹水），以及它们的潜力，作为疾病的指标，扩大了兴趣，并加强了微泡生物学和功能的研究工作。本申请的首要目标是确定微泡形成的分子机制。该项目的重点是中心假设，即特定的ARF和Rab蛋白直接膜型蛋白酶和其他蛋白质的微泡生物发生的网站和RhoA和Rac1信号之间的紧密交换控制微泡和侵袭介导的蛋白水解入侵之间的切换所需的可塑性。我们将讨论两个具体目标。在第一个目标中，我们将定义内吞再循环途径，直接货物的微泡生物发生的网站，以及研究如何招聘特定的Rab效应调节肌动球蛋白为基础的收缩所需的微泡生物发生。在第二个目标中，我们将研究侵袭性肿瘤细胞中RhoA和Rac 1的空间激活。我们还将研究在微泡形成过程中调节Rac 1下调的潜在机制，以及Rho信号如何促进这一过程。鉴于最近对微泡的生物学和临床前景的高度兴趣，这些研究是非常流行的。它们将推进目前对微泡生物发生的理解，并有可能为诊断和治疗应用提供靶点。

DESCRIPTION (provided by applicant): This is an application to investigate the mechanisms of microvesicle biogenesis in invasive tumor cells. It builds on exciting findings generated with previous NIH funding on a unique population of vesicles, called microvesicles that contain functionally active proteases and are released by tumor cells as they acquire invasive potential. The release of protease-loaded microvesicles may serve as a mechanism to bring about matrix degradation and perhaps even deposit paracrine information at distal locations, thus creating paths of "least resistance" as tumor cells invade and migrate through surrounding tissue. This is distinct from pericellular proteolysis at invadopodia, which enables localized matrix degradation juxtaposed to the leading edge. Discovering that there may exist more than one mode of proteolytic invasion, limits the effectiveness of any invasion-targeted therapeutic strategy that does not include both focal and distal proteolysis. While a significant amount of research has been directed to the understanding mechanisms of invadopodia formation and function at sites of cell invasion, microvesicles biogenesis and function remains a relatively understudied area of tumor biology. However, recent accruing evidence demonstrating the bona fide presence of microvesicles in body fluids (blood, urine and ascites), and their potential to serve as indicators of disease, has extended interest and intensified research efforts in microvesicle biology and function. The overarching objective of this application is to define molecular mechanisms of microvesicle formation. The project focuses on the central hypothesis that specific ARF and Rab proteins direct membrane type proteases and other proteins to sites of microvesicle biogenesis and that tight interchanges between RhoA and Rac1 signaling governs the plasticity required for switching between microvesicle and invadopodia-mediated proteolytic invasion. We will address two specific aims. In the first aim, we will define endocytic recycling pathways that direct cargo to sites of microvesicle biogenesis as well as examine how recruitment of specific Rab effectors regulate actomyosin-based contraction required for microvesicle biogenesis. In the second aim, we will examine the spatial activation of RhoA and Rac1 in invasive tumor cells. We will also investigate potential mechanisms that regulate Rac1 down regulation during microvesicle formation and how Rho signaling facilitates the process. Given recent heightened interest in the biology and clinical promise of microvesicles, these investigations are highly current. They will advance present understanding of microvesicle biogenesis and have potential to provide targets for diagnostic as well as therapeutic application.