PI3K opposes p27 and G1 arrest by TGF-B in human cancer

描述（由申请人提供）：转化生长因子- β (tgf - β)通过激活cdk抑制剂介导细胞周期阻滞，其中的原理是p27[Kip1]。tgf - β的生长抑制作用的丧失发生在肿瘤发生的早期，并有助于肿瘤的进展。我们已经研究了tgf - β的细胞周期效应以及它们如何在人类肿瘤中丢失，由于p27蛋白水解加速或p27细胞质错定位，p27在人类癌症中经常失活。在癌细胞中，渐进性检查点丢失使得p27对于tgf - β介导的阻滞至关重要。我们发现磷酸化肌醇3’激酶（PI3K）效应物激活后，蛋白激酶B （PKB）在苏氨酸157位点磷酸化p27，使p27在细胞质中定位错误。PKB激活的细胞也显示细胞周期蛋白D1-cdk复合物中p27的增加。初步数据表明，PI3K通路的其他效应物也可能抑制p27的功能。在癌症中，PI3K及其下游效应通路通常通过致癌受体酪氨酸激酶过表达、激活ras突变或PI3K抑制剂PTEN的缺失而被激活。在这里，我们研究了p27功能和G1被tgf - β阻滞是如何在癌症中被有丝分裂PI3K信号所反对的。我们的假设是，PI3K通路的组成性激活导致p27磷酸化事件，从而降低其对细胞周期蛋白E-cdk2的亲和力，改变其稳定性和细胞内定位，并取消其抑制功能，从而导致tgf - β抗性。目的1)检测PKB是否激活p27对细胞周期蛋白D1-cdk4的组装功能，并降低p27对细胞周期蛋白E-cdk2的抑制作用；AIM 2)确定PKB以外的PI3K效应物是否介导tgf - β耐药；AIM 3)检测其他PI3K依赖性激酶是否能磷酸化p27并改变其功能；AIM 4)研究PI3K效应物是否影响p27的核输入。这些研究可能将tgf - β抵抗与人类癌症中常见的有丝分裂P13K信号通路的致癌激活联系起来。阐明肿瘤中p27失活和tgf - β耐药的分子机制可能为治疗干预提供新的靶点。

DESCRIPTION (provided by applicant): Transforming growth factor-beta (TGF-beta) mediates cell cycle arrest by activating cdk inhibitors, principle among which is p27[Kip1]. Loss of growth inhibition by TGF-beta occurs early in oncogenesis and contributes to tumor progression. We have studied cell cycle effects of TGF-beta and how they are lost in human tumors, p27 is often inactivated in human cancers due to accelerated p27 proteolysis or cytoplasmic mislocalization of p27. In cancer cells, progressive checkpoint loss makes p27 essential for TGF-beta mediated arrest. We showed activation of the phosphoinositol 3' kinase (PI3K) effector, protein kinase B (PKB) phosphorylates p27 at threonine 157, mislocalizating p27 in the cytoplasm. PKB activated cells also showed increased p27 in cyclin D1-cdk complexes. Preliminary data suggest that other effectors of the PI3K pathway may also inhibit p27 function. PI3K and its downstream effector pathways are often activated in cancers by oncogenic receptor tyrosine kinase overexpression, activating ras mutations or loss of the PI3K inhibitor, PTEN. Here we investigate how p27 function and G1 arrest by TGF-beta may be opposed by mitogenic PI3K signaling in cancers. Our hypothesis is that constitutive activation of the PI3K pathway leads to p27 phosphorylation events that reduce its affinity for cyclin E-cdk2, alter its stability and intraceltular localization, and abrogate its inhibitory function leading to TGF-beta resistance. This is pursued by the following specific aims: AIM 1) to assay if PKB activates p27 assembly function toward cyclin D1-cdk4 and reduces p27's inhibitory action toward cyclin E-cdk2; AIM 2) to determine whether PI3K effectors other than PKB can mediate TGF-beta resistance; AIM 3) to assay if other PI3K dependent kinases can phosphorylate p27 and alter its function and AIM 4) to investigate whether PI3K effectors impair nuclear import of p27. These studies may link TGF-beta resistance to oncogenic activation of mitogenic P13K signaling pathways commonly observed in human cancers. Elucidation of molecular mechanisms of p27 inactivation and TGF-beta resistance in cancers may yield new targets for therapeutic intervention.