Molecular Mechanisms of Cell Invasion

这是一个研究肿瘤细胞侵袭的分子机制的建议。习得 侵袭潜力赋予肿瘤细胞从原发肿瘤分离并扩散到远端的能力 网站.使用侵袭性黑色素瘤细胞系作为模型系统，我们已经发现， ARF 6蛋白调节细胞侵袭。近年来，ARF 6已成为一种重要的信号， 分子，并已显示在多种细胞类型中调节内体循环和肌动蛋白重塑， 这两者都可能影响细胞迁移/侵入潜能的获得。我们的研究表明 ARF 6的激活通过一个信号通路与ERK的激活有关， for cell细胞invasion入侵.通过siRNA沉默ARF 6或表达ARF 6显性阴性突变体 在基于细胞的体外测定中消除了侵袭。我们还发现内源性ARF 6的激活 增加对刺激肿瘤细胞侵袭的生理刺激的反应。在本申请中，我们的目标是 研究ARF 6调节细胞侵袭的机制。根据初步调查结果，我们 假设ARF 6激活至少部分通过调节（1） 金属蛋白酶的运输和（2）侵袭伪足形成所需的细胞骨架重塑。通过使用 细胞生物学，分子和生物化学方法，细胞和动物模型系统，我们将 研究ARF 6在肿瘤细胞侵袭中的作用。我们提出三个具体目标。在目标1中， 通过检测ARF 6对MT 1-MMP的调节，研究其在蛋白酶运输调节中的作用 运输和从细胞表面释放蛋白酶。在目的2中，我们将阐明信号转导 细胞侵袭过程中ARF 6与Rac 1活化的连接途径，并询问侵袭伪足的形成是否重要 用于蛋白酶分泌。在目的3中，我们将研究ARF 6 GT3循环在黑色素瘤细胞中的作用。 使用小鼠模型系统进行体内侵袭。我们相信从调查中获得的信息 将为蛋白酶运输的调节提供新的见解，这是一个相对研究不足的癌症领域。 生物学，并将扩大现有的知识，在肿瘤进展过程中发生的分子变化。 希望这些发现将最终有助于设计新的诊断和/或治疗策略。 癌症进展的治疗干预。

This is a proposal to investigate the molecular mechanisms underlying tumor cell invasion. Acquistion of invasive potential confers on tumor cells the ability to detach from the primary tumor and spread to distal sites. Using an invasive melanoma cell line as a model system, we have found that the small GTP-binding protein, ARF6, regulates cell invasion. In recent years, ARF6 has emerged as an important signaling molecule and has been shown to regulate endosome cycling, and actin remodeling in a variety of cell types, both of which can impinge on the acquisition of cell migratory/invasive potential. Our studies demonstrate that ARF6 activation is linked to the activation of ERK via a signaling pathway that appears to be essential for cell invasion. Silencing of ARF6 by siRNAs or expression of an ARF6 dominant negative mutant abolishes invasion in cell-based in vitro assays. We have also shown that activation of endogenous ARF6 increases in response to physiological stimuli that stimulate tumor cell invasion. In this application, we aim to investigate the mechanisms by which ARF6 regulates cell invasion. Based on preliminary findings, we hypothesize that ARF6 activation promotes tumor cell invasion at least in part, by regulating (1) the trafficking of metalloproteases and (2) cytoskeletal remodeling required for invadopodia formation. By using cell biological, molecular and biochemical approaches in both, cell and animal model systems, we will investigate the role of ARF6 in tumor cell invasion. We propose three specific aims. In aim 1, we will investigate the role of ARF6 in the regulation of protease traffic by examining its regulation of MT1-MMP traffic and the release of proteases from the cell surface. In aim 2, we will elucidate the signal transduction pathway linking ARF6 to Rac1 activation during cell invasion and ask if invadopodia formation is important for protease secretion. In aim 3, we will investigate the role of the ARF6 GTPase cycle in melanoma cell invasion in vivo using a mouse model system. We believe that the information gained from investigations will provide new insights into the regulation of protease trafficking, a relatively understudied area of cancer biology, and will broaden existing knowledge on the molecular changes that occur during tumor progression. It is hoped that these findings will ultimately aid in the design of new strategies for diagnostic and/or therapeutic intervention of cancer progression.