REGULATIONS OF BETA CATENIN BY PROTEIN PHOSPHATASE 2A

对结直肠癌的分析表明，Wnt通路的失控是结直肠癌恶变的关键步骤。因此，了解这一途径的调控可能会改善人类癌症的预防和治疗。Wnt途径在结肠中的一个重要功能是通过磷酸化调节的信号转导途径调节β-连环素依赖的转录。虽然已有几种激酶被证明调节Wnt途径，但先前没有明确地确定磷酸酶是该途径的一个组成部分。我们的数据表明，蛋白磷酸酶2A(PP2A)的B56调节亚基与腺瘤性息肉病结肠(APC)蛋白相互作用，B56的过度表达导致β-连环素丰度降低，导致哺乳动物组织培养细胞和非洲爪哇胚胎外植体中β-连环素依赖基因的表达减少。B56的过表达似乎激活了蛋白酶体介导的磷酸化依赖的β-连环蛋白的降解。此外，磷酸酶抑制剂冈田酸导致β-连环蛋白水平增加，如果PP2A抑制Wnt信号转导，这一点是可以预期的。因此，我们的结果表明，含有B56的PP2A全酶是Wnt信号转导途径的关键调节因子。本研究的目的是明确B56和PP2A在Wnt信号转导通路和肠癌发生发展中的确切作用。我们建议(1)确定作为PP2A：B56底物的Wnt途径的关键成分，(2)确定途径B56的哪些额外成分相互作用，(3)测试Wnt是否部分通过调节PP2A：B56的活性来传播其信号，以及(4)使用转基因小鼠来检验B56的过度表达抑制结肠癌发生，而显性阴性的B56等位基因促进其发生的假设。

The analysis of colorectal carcinomas suggests that deregulation of the Wnt pathway is an essential step in their malignant transformation. Understanding the regulation of this pathway may therefore lead to improved prevention and treatment of human carcinomas. One crucial function of the Wnt pathway in the colon is to regulate beta-catenin-dependent transcription via a phosphorylation-regulated signal transduction pathway. Although several kinases have been shown to regulate the Wnt pathway, no phosphatase has previously been clearly identified as a component of this pathway. Our data show that the B56 regulatory subunits of protein phosphatase 2A (PP2A) interact with the adenomatous polyposis coli (APC) protein, and that B56 overexpression causes decreased beta-catenin abundance, leading to decreased beta-catenin-dependent gene expression in mammalian tissue culture cells and Xenopus embryo explants. The overexpression of B56 appears to activate the phosphorylation-dependent, proteasome-mediated, degradation of beta-catenin. In addition, the phosphatase inhibitor okadaic acid causes increased levels of beta-catenin, as would be expected if PP2A inhibited Wnt signaling. Our results suggest, therefore, that the B56-containing PP2A holoenzyme is a key regulator of a Wnt signal transduction pathway. The objective of this proposal is to define the precise role of B56 and PP2A in the Wnt signal transduction pathway and the development of intestinal cancers. We propose to (1) identify critical Wnt pathway components that are PP2A:B56 substrates, (2) determine with which additional components of the pathway B56 interacts, (3) test if Wnt propagates its signal in part by regulating PP2A:B56 activity, and (4) use transgenic mice to test the hypothesis that overexpression of B56 suppresses colon carcinogenesis, while dominant-negative B56 alleles promote it.