ROLE OF CYTOPLASMIC P27KIP1 IN CELL MOTILITY AND METASTASIS

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。转移性肿瘤生长是导致癌症患者死亡的主要事件，然而，其分子基础目前尚不清楚。转移性肿瘤细胞通常比正常细胞或非转移性肿瘤细胞迁移速度更快。这一观察结果导致了一种假设，即细胞运动(迁移)的增加是细胞发生转移所必需的。最近，一个新的细胞周期蛋白依赖性作用：CDK抑制物p27KIP1在细胞运动中被发现。结果表明，HGF信号通过Met受体导致p27KIP1的核输出进入细胞质。这种胞浆定位是p27KIP1诱导细胞运动所必需的，并导致肌动蛋白细胞骨架重排。这些结果表明，肿瘤细胞中的信号失控诱导了p27的胞浆输入。我们的中心假设是，在细胞质中，p27组装成一个参与细胞骨架重塑和细胞迁移的复合体，导致转移增加。P27在细胞质中的重新定位和参与肌动蛋白细胞骨架重排，在进化上使人联想到酵母中的阿尔法因子信息素信号。酵母细胞周期蛋白：CDK抑制剂Far1p既介导了细胞核中G1期细胞周期的停滞，又诱导了细胞质中shmoo的形成，使肌动蛋白细胞骨架朝向相反的交配伙伴。作为对α因子的反应，Far1p从胞核移位到细胞质，并与CDc24p相互作用，CDc24p是一种与CDc42p的鸟核苷酸交换因子。与这一功能相似的是，存在于p27散发域的Far1p样序列基序的突变使细胞迁移和肌动蛋白重排失活，但保留了p27的细胞周期停滞功能。我们的目标是通过TAP-Tag纯化和质谱分析来鉴定细胞质中与p27结合的蛋白质复合体。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Metastasic tumor growth is the principal event leading to death in patients with cancer, however, its molecular basis is presently poorly understood. Metastasic tumor cells generally migrate at a faster rate than normal cells or non-metastasic tumor cells. This observation leads to the hypothesis that increased cell motility (migration) is necessary for cells to become metastasic. Recently, a novel cytosolic-dependent role for the cyclin:CDK inhibitor p27KIP1 in cell motility was identified. It was shown that HGF signaling via the Met receptor resulted in the nuclear export of p27 KIP1 into the cytoplasm. This cytosolic localization was required for p27 KIP1 to induce cell motility and resulted in actin cytoskeletal rearrangements. These results suggest that deregulated signaling in tumor cells induce cytoplasmic import of p27. Our central hypothesis is that in the cytoplasm, p27 assembles into a complex involved in cytoskeletal remodeling and cell migration resulting in increased metastasis. The cytoplasmic relocalization and involvement of p27 in actin cytoskeletal rearrangement are evolutionarily reminiscent of alpha factor pheromone signaling in yeast. Far1p, the yeast cyclin:cdk inhibitor, mediates both a G1 cell cycle arrest in the nucleus and induction of shmoo formation in the cytoplasm that orients the actin cytoskeleton toward the opposite mating partner . In response to alpha factor, Far1p translocates from the nucleus to the cytoplasm and interacts with Cdc24p, a guanine nucleotide exchange factor for Cdc42p. Consistent with this functional analogy, mutation of a Far1p-like sequence motif present in the p27 scatter domain inactivates both cell migration and actin rearrangement, but preserves the cell cycle arrest functions of p27. Our objective is to identify the proteins complexes binding to p27 in the cytoplasm by TAP-tag purification and mass spectrometry analysis.