C Cbl in Modulation of Cell Adhesion and Morphology

C Cbl 调节细胞粘附和形态

• 批准号: 6757419
• 负责人: ALEXANDER Y TSYGANKOV
• 金额: $ 1.32万
• 依托单位: TEMPLE UNIV OF THE COMMONWEALTH
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-07-01 至 2006-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6757419
• 关键词: athymic mouse; biological signal transduction; cell adhesion; connective tissue neoplasm; enzyme activity; extracellular matrix proteins; fibroblasts; guanosinetriphosphatases; interleukin 3; oncoproteins; protein tyrosine kinase; ubiquitin; viral carcinogenesis; virus related neoplasm /cancer

DESCRIPTION: (Adapted from the investigator's abstract) The protooncogenic protein c-Cbl is tyrosine phosphorylated both in normal cells in response to various stimuli and in cells transformed by oncogenic protein tyrosine kinases (PTKs). Tyrosine phosphorylation of c-Cbl enhances its binding to crucial signaling proteins. c-Cbl also contains an SH2-like domain capable of binding to several PTKs and a large number of proline-rich motifs capable of binding to multiple SH3 domain-containing proteins. The ability of c-Cbl to interact with a wide variety o signaling proteins argues in favor of c-Cbl acting as a multivalent adaptor protein. Furthermore, c-Cbl is capable of inhibiting activities of some PTKs. This inhibition may be explained, at least partially, by the recently discovered ability of c-Cbl to facilitate ubiquitination of Cbl-associated PTKs, In spite of the abundance of biochemical data, biological functions of c-Cbl remain poorly understood. In particular, it remains unclear whether c-Cb1 can positively regulate cell functions. Furthermore, effects of protooncogenic c-Cb1 on cell transformation have not been characterized in detail. We have been studying tyrosine phosphorylation of c-Cb1 and its functional role in cell activation and transformation. Our analysis of biological functions of c-Cbl in v-Abl-transformed NIH 3T3 fibroblasts demonstrated that wild-type c-Cbl, but not its tyrosine phosphorylation-defective mutants, facilitates adhesion and spreading of transformed cells and reduces their anchorage independence, exhibiting an overall transformation-suppressing effect. We have recently shown that the c-Cb1-mediated reversion of morphological transformation of these cells is caused by an increase in extracellular matrix production, and that this increase is linked to the c-Cbl-dependent activation of small GTPases regulating cytoskeletal rearrangements. The current proposal is designed to further develop this research. Its overall objective is to understand the role of c-Cb1 in the regulation of cell adhesion and morphology and to determine the molecular basis of these effects. The hypothesis to be tested in the proposed study is that these biological effects of c-Cbl are mediated by triggering of P1-3' kinase, Vav2 and Crk-dependent signaling leading to activation of Rho-family GTPases and, possibly, Rap 1. We further hypothesize that ubiquitination-driven degradation of proteins interfering with the assembly offocal adhesions and stress fibers is involved in the effects of c-Cbl on v-Abl-transformed cells. Accordingly, the specific aims of our project are as follows: 1. To determine relative contributions of Rho-like and Rap GTPases to the observed c-Cbl-dependent facilitation of adhesion and spreading of v-Abl-transformed fibroblasts. 2. To elucidate the mechanisms of activation of small GTPases involved in c-Cbl-dependent facilitation of adhesion and spreading of v-Abl-transformed fibroblasts and to assess relative contributions of these mechanisms to the overall biological effect of c-Cbl. 3. To elucidate the mechanisms whereby c-Cbl-dependent ubiquitination is involved in the effects of c-Cbl on v-Abl-transformed fibroblasts. 4. To determine the effect of c-Cbl overexpression on adhesion and transformation potential of hematopoietic cells transformed with Bcr-Abl, a constitutively active PTK, which causes chronic myeologenous leukemia.

描述：(改编自研究人员的摘要)原癌基因 蛋白c-Cbl在正常细胞中都被酪氨酸磷酸化，以响应 不同刺激和致癌蛋白酪氨酸激酶转化的细胞内 (PTK)。C-Cbl酪氨酸磷酸化增强其与关键蛋白的结合 信号蛋白。C-Cb1还含有能够结合的SH2样结构域 与几个PTK和大量富含Pro的基序能够结合 多个含SH3结构域的蛋白质。C-Cbl与Cbl相互作用的能力 各种各样的信号蛋白都支持c-Cbl作为一种 多价适配子蛋白。此外，c-Cbl能够抑制 一些PTK的活动。这种抑制可以解释，至少部分地， 通过最近发现的c-Cbl促进泛素化的能力 CBL相关的PTKs，尽管有丰富的生化数据，但生物学上 C-Cbl的功能仍然知之甚少。特别是，目前仍不清楚 C-CB1是否能正向调节细胞功能。此外， 原癌基因c-CB1在细胞转化中的作用尚未被表征 细节。 我们一直在研究c-cb1的酪氨酸磷酸化及其功能作用。 在细胞的激活和转化中。我们对细胞生物学功能的分析 在v-Abl转化的NIH3T3成纤维细胞中，c-Cbl证明野生型 C-Cb1，但不是它的酪氨酸磷酸化缺陷突变体，促进了 转化细胞的黏附和扩散，并减少其锚定 独立性，表现出一种整体的转型抑制效应。我们有 最近发现c-CB1介导的形态逆转 这些细胞的转化是由细胞外基质的增加引起的 这种增加与c-Cbl依赖的激活有关 调节细胞骨架重排的小GTP酶。 目前的建议旨在进一步发展这项研究。它的整体 目的是了解c-cb1在细胞黏附调节中的作用。 和形态，并确定这些效应的分子基础。这个 在拟议的研究中要检验的假设是，这些生物效应 C-Cbl的表达是通过触发依赖于P1一3‘的蛋白、Vav2和Crk来实现的 信号导致Rho家族GTP酶的激活，可能还有Rap 1。 进一步假设泛素化驱动的蛋白质降解 干扰局部粘连和应力纤维的组装 C-Cbl对v-Abl转化细胞的影响。相应地，具体的 我们项目的目标如下：1.确定相对贡献 Rho样和Rap GTP酶对c-Cbl依赖的易化作用的影响 V-Abl转化成纤维细胞的黏附和铺展。 2.阐明小分子GTP酶激活的机制。 C-Cbl对v-Abl转化细胞黏附和铺展的促进作用 成纤维细胞，并评估这些机制对 C-Cbl的整体生物学效应。 3.阐明c-Cbl依赖泛素化的机制 参与c-Cbl对v-Abl转化成纤维细胞的影响。 4.检测c-Cbl过表达对细胞黏附和黏附的影响 Bcr-Abl、a转化造血细胞的转化潜能 体质活性的PTK，导致慢性粒细胞白血病。