Regulation of a lipid raft trafficking pathway by integrins

描述（由申请人提供）：整合素介导的粘附控制生长因子和细胞因子受体下游许多信号的传递。这些作用调节细胞生长、分化和许多其他功能。我们之前的工作确定了整合素调节的膜运输途径，介导了许多这些影响。锚定依赖的细胞脱离细胞外基质触发脂筏成分通过小泡内化。这些结构域在质膜上的清除抑制了Rho家族gtpase、Erk和Akt的生长因子激活。在细胞外基质上复制细胞触发这些成分的胞吐，促进细胞扩散和恢复信号。在上一个资助期间，我们确定了Arf6是胞吐作用第一步的关键决定因素：从循环内体中退出筏成分。然后，我们发现RalA通过与囊泡的相互作用介导了下一步，即胞囊囊在质膜上的对接。在许多系统中，RalA被认为是生长和癌症转移的锚定依赖性的特异性调节因子，因此，我们的数据链接整合素调节肿瘤生长和转移的膜运输。我们现在建议研究整合素控制胞吐途径的机制，并研究其在细胞运动中的作用。具体而言，我们将通过鉴定介导整合素激活RalA的相关鸟嘌呤核苷酸交换因子，阐明整合素在脂筏胞吐过程中调控RalA的机制，并进一步明确其激活机制。我们将同样确定介导Arf6激活的交换因子并研究其机制。最后，我们将阐明整合素调节的脂筏运输在细胞运动中的作用，特别是是否存在通过相同途径的脂筏运输的空间限制调节以及它如何促进定向细胞运动。这项工作将有助于我们理解细胞行为的基础以及调节癌症转移的关键途径。

DESCRIPTION (provided by applicant): Integrin-mediated adhesion controls the transmission of many signals downstream of growth factor and cytokine receptors. These effects modulate cell growth, differentiation and many other functions. Our previous work identified an integrin-regulated membrane trafficking pathway that mediates a number of these effects. Detachment of anchorage- dependent cells from their extracellular matrix triggers internalization of lipid raft components via caveolae. The resultant clearance of these domains from the plasma membrane inhibits growth factor activation of Rho family GTPases, Erk and Akt. Replating cells on extracellular matrix triggers exocytosis of these components to promote cell spreading and restore signaling. During the last grant period, we identified Arf6 as the key determinant of the first step of exocytosis: exit of raft components from the recycling endosomes. We then showed that RalA mediates the next step, docking of exocytic vesicles at the plasma membrane through its interaction with the exocyst. RalA has been implicated as a specific regulator of anchorage-dependence of growth and cancer metastasis in many systems, thus, our data link integrin regulation of membrane trafficking with tumor growth and metastasis. We now propose to investigate the mechanisms by which integrins control the exocytosis pathway and investigate its role in cell motility. Specifically, we will elucidate the mechanism by which integrins regulate RalA during lipid raft exocytosis by identifying the relevant guanine nucleotide exchange factor that mediates integrin activation of RalA and further define the mechanism of activation. We will similarly identify the exchange factors that mediate Arf6 activation and investigate mechanism. Lastly, we will elucidate the role of integrin-regulated lipid raft transport in cell motility, specifically whether there is spatially restricted regulation of lipid raft trafficking via the same pathway and how it contributes to directed cell movement. This work will contribute to our understanding of a pathway that is fundamental to cell behavior as well as crucial in regulating cancer metastasis.