REGULATION OF PROTEIN PHOSPHATASE 2A BY CELLULAR PROTEIN

本申请的长期目标是表征似乎是调节蛋白磷酸酶2A（PP 2A）活性的新细胞机制，PP 2A是一种通过使这些途径中的关键蛋白脱磷酸化而在调节细胞生长调节途径中起关键作用的酶。具体而言，我们建议，一种称为HSF 2的蛋白质调节PP 2A的活性结合到PP 2A的PR 65/A亚基，并通过直接与催化亚基竞争其在PR 65中的结合位点来防止其与催化亚基的关联。HSF 2阻断PR 65和催化亚基之间相互作用的能力的生物学相关性通过显示PR 65中阻止催化亚基结合的突变与人肺癌和结肠癌相关的结果来证明。HSF 2也是已知的是一种转录因子，其调节热休克蛋白（hsp）基因的表达，因此，我们假设，HSF 2的PP 2A的调节可以作为一种机制之间的串扰控制的hsp表达和细胞生长调节途径。为了阐明HSF 2作为PP 2A的新型调节剂的功能，我们提出1）鉴定HSF 2和PR 65中对于这种相互作用重要的序列，2）表征调节HSF 2：PR 65相互作用的细胞机制，3）确定HSF 2在调节细胞PP 2A活性中的功能，以及4）使用酵母双杂交相互作用筛选鉴定这种新型PP 2A调节蛋白的其他潜在成员（我们已经使用这种方法鉴定了一种新的候选物，称为mel-18的生长调节蛋白）。这些研究将通过阐明PP 2A调节的新机制以及细胞中两个重要信号通路之间的串扰的新机制来推进知识，热休克蛋白调节和细胞生长/分化的控制。这些知识可以帮助理解正常细胞生长调节以及癌细胞中的异常增殖控制。

The long-term objective of this application is to characterize what appears to be a new cellular mechanism for regulating the activity of protein phosphatase 2A (PP2A), an enzyme which plays a critical role in modulating cell growth regulatory pathways by dephosphorylating key proteins in these pathways. Specifically, we propose that a protein called HSF2 regulates PP2A activity by binding to the PR65/A subunit of PP2A and preventing its association with the catalytic subunit by directly competing with catalytic subunit for its binding site in PR65. The biological relevance of HSF2's ability to block interaction between PR65 and catalytic subunit is demonstrated by results showing that mutations in PR65 that prevent binding of catalytic subunit are associated with human lung and colon cancers. HSF2 is also known to be a transcription factor which regulates expression of heat shock protein (hsp) genes, and thus we hypothesize that HSF2 regulation of PP2A could serve as a mechanism for cross-talk between control of hsp expression and cell growth regulatory pathways. To elucidate the function of HSF2 as a novel regulator of PP2A we propose to 1) identify the sequences in HSF2 and PR65 important for this interaction, 2) characterize cellular mechanisms which regulate the HSF2:PR65 interaction, 3) determine the function of HSF2 in regulating cellular PP2A activity, and 4) identify other potential members of this new class of PP2A regulatory protein using yeast two-hybrid interaction screens (we have already identified one new candidate using this meth6d, a growth regulatory protein called mel-18). These studies will advance knowledge by elucidating a new mechanism of PP2A regulation as well as a new mechanism for cross-talk between two important signaling pathways in cells, hsp regulation and control of cell growth/differentiation. This knowledge could aid understanding of both normal cell growth regulation as well as aberrant proliferation control in cancer cells.