REGULATIONS OF BETA CATENIN BY PROTEIN PHOSPHATASE 2A

结直肠癌的分析表明，Wnt通路的失调是其恶性转化的重要步骤。 因此，了解这一途径的调节可能会改善人类癌症的预防和治疗。 结肠中Wnt通路的一个关键功能是通过磷酸化调节的信号转导通路调节β-连环蛋白依赖性转录。 虽然已经显示了几种激酶调节Wnt途径，但以前没有磷酸酶被明确鉴定为该途径的组分。我们的数据表明，蛋白磷酸酶2A（PP 2A）的B56调节亚基与腺瘤性结肠息肉病（APC）蛋白相互作用，B56过表达导致β-连环蛋白丰度降低，导致哺乳动物组织培养细胞和爪蟾胚胎外植体中β-连环蛋白依赖性基因表达降低。 B56的过表达似乎激活了磷酸化依赖的、蛋白酶体介导的β-连环蛋白降解。 此外，磷酸酶抑制剂冈田酸导致β-连环蛋白水平增加，如PP 2A抑制Wnt信号传导所预期的那样。因此，我们的研究结果表明，含有B56的PP 2A全酶是Wnt信号转导途径的关键调节因子。 该提案的目的是确定B56和PP 2A在Wnt信号转导途径和肠癌发展中的确切作用。 我们建议（1）确定Wnt途径的关键成分是PP 2A：B56底物，（2）确定与途径B56的其他成分相互作用，（3）测试Wnt是否部分通过调节PP 2A：B56活性来传播其信号，（4）使用转基因小鼠来测试B56过表达抑制结肠癌发生，而显性阴性B56等位基因促进结肠癌发生的假设。

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.