The Role of MIM in growth factor signaling and cell motility

MIM 在生长因子信号传导和细胞运动中的作用

• 批准号: 7483189
• 负责人: Steven ZHAN
• 金额: $ 25.59万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7483189
• 关键词: Achievement; Actins; Area; Binding; Biochemical; Bladder; Cell Adhesion; Cell Surface Extensions; Cells; Cellular Structures; Cicatrix; Classification; Complex; Cytoskeleton; Data; Development; Distant; Family; Future; Gene Expression; Goals; Growth Factor; Human; In Vitro; Intervention; Invasive; Knock-out; Lead; Length; Malignant Epithelial Cell; Mediating; Membrane; Molecular; Monomeric GTP-Binding Proteins; Mutation; Neoplasm Metastasis; Normal Cell; Organ; Pathway interactions; Phosphorylation; Phosphotransferases; Platelet-Derived Growth Factor; Primary Lesion; Protein Family; Protein Overexpression; Protein Tyrosine Kinase; Proteins; RNA Interference; Regulation; Role; Signal Pathway; Signal Transduction; Temperature; Testing; Tyrosine; Tyrosine Phosphorylation; base; bladder Carcinoma; cancer cell; cell motility; design; human EMS1 protein; in vivo; member; migration; neoplastic cell; novel; polymerization; response; rho; rho GTP-Binding Proteins; src-Family Kinases; tumor; tumor progression; v-src Oncogenes

DESCRIPTION (provided by applicant): Tumor spreading occurs as a consequence of continuous cell membrane protrusion, invasion and migration, the activities that require dynamic actin cytoskeleton reorganization. Actin assembly in the cell leading edge is achieved by the functions of Arp2/3 complex, WASP/WAVE family proteins and cortactin, which are activated by Rho small GTPases and protein tyrosine kinase Src. Our recent study has uncovered that cortactin binds to the protein product of missing in metastasis (MIM) gene, expression of which is frequently lost in a subset of aggressive tumor cells and advanced human bladder cell carcinomas. Overexpression of MIM inhibits markedly the motility of tumor cells or normal cells mediated by growth factors, and restrains the lamellipodia formation induced by constitutively activated Rac1, a member of the Rho GTPase family. Within cells MIM colocalizes with and is able to coprecipitate with Rac1. In vitro, MIM inhibits the actin polymerization mediated by WASP. Interestingly, MIM undergoes a rapid tyrosine phosphorylation in concurrent with transient interaction with cortactin in response to growth factors. Phosphorylation at two tyrosine residues is required for MIM to inhibit Rac1. Based on these data, we hypothesize that MIM is implicated in a novel signaling pathway antagonizing the function of Rac in the assembly of actin cytoskeleton, the activity that is subjected to a regulation through tyrosine phosphorylation. To test this hypothesis, we propose to characterize the mechanism for MIM to antagonize Rac. Therefore, we will determine whether MIM interacts directly with Rac and examine whether the direct association is necessary and sufficient to inhibit Rac. We will also analyze the role of IRSp53 and WAVE, the effectors of Rac, in the function of MIM, and test whether inhibition of Rac is the mechanism for MIM to modulate cell motility induced by growth factors. We will delineate the signaling pathway for MIM phosphorylation and examine whether Src is the primary tyrosine kinase responsible for the PDGF mediated tyrosine phosphorylation of MIM, and analyze tyrosine phosphorylation of MIM in Src knockout cells and the cells overexpressing temperature sensitive v-Src. We will also explore the role of tyrosine phosphorylation in the regulation of MIM with respect to interaction with Rac and cortactin, and cell motility. Finally, we will examine the role of MIM expression at low levels in the motility of tumor cells by silencing MIM using RNA interference in MIM expression cells, and examine whether suppression of MIM expression will lead to increase in lamellipodia development, Rac activation, actin polymerization in cells and cell motility. It is anticipated that achievement of the goal of this proposal will direct our future effort to understand how MIM contributes to tumor progression.

描述（由申请人提供）：肿瘤扩散是由于连续的细胞膜突出、侵袭和迁移而发生的，这些活动需要动态肌动蛋白细胞骨架重组。细胞前沿的肌动蛋白组装是通过 Arp2/3 复合物、WASP/WAVE 家族蛋白和皮质蛋白的功能实现的，这些蛋白由 Rho 小 GTP 酶和蛋白酪氨酸激酶 Src 激活。我们最近的研究发现，cortactin 与转移缺失 (MIM) 基因的蛋白质产物结合，该基因的表达在侵袭性肿瘤细胞和晚期人类膀胱细胞癌的亚群中经常丢失。 MIM的过度表达显着抑制生长因子介导的肿瘤细胞或正常细胞的运动，并抑制由组成型激活的Rac1（Rho GTPase家族成员）诱导的板状伪足形成。在细胞内，MIM 与 Rac1 共定位并能够与其共沉淀。在体外，MIM 抑制 WASP 介导的肌动蛋白聚合。有趣的是，MIM 经历快速酪氨酸磷酸化，同时响应生长因子与 Cortactin 短暂相互作用。 MIM 需要两个酪氨酸残基磷酸化才能抑制 Rac1。基于这些数据，我们假设 MIM 涉及一种新的信号通路，该通路拮抗肌动蛋白细胞骨架组装中 Rac 的功能，该活性通过酪氨酸磷酸化受到调节。为了检验这一假设，我们建议描述 MIM 拮抗 Rac 的机制。因此，我们将确定MIM是否与Rac直接相互作用，并检验这种直接关联对于抑制Rac是否是必要和充分的。我们还将分析 Rac 的效应子 IRSp53 和 WAVE 在 MIM 功能中的作用，并测试抑制 Rac 是否是 MIM 调节生长因子诱导的细胞运动的机制。我们将描绘MIM磷酸化的信号通路，并检查Src是否是负责PDGF介导的MIM酪氨酸磷酸化的主要酪氨酸激酶，并分析Src敲除细胞和过表达温度敏感v-Src的细胞中MIM的酪氨酸磷酸化。我们还将探讨酪氨酸磷酸化在 MIM 调节中的作用，即与 Rac 和 cortactin 的相互作用以及细胞运动。最后，我们将通过在MIM表达细胞中使用RNA干扰来沉默MIM来检查低水平的MIM表达在肿瘤细胞运动中的作用，并检查抑制MIM表达是否会导致板状伪足发育、Rac激活、细胞内肌动蛋白聚合和细胞运动的增加。预计该提案目标的实现将指导我们未来努力了解 MIM 如何促进肿瘤进展。