REGULATION OF PROTEIN PHOSPHATASE 2A BY CELLULAR PROTEIN

这一应用的长期目标是描述一种似乎是调节蛋白磷酸酶2A(PP2A)活性的新的细胞机制，PP2A是一种通过去磷酸化这些途径中的关键蛋白而在调节细胞生长调节途径中发挥关键作用的酶。具体地说，我们认为一种名为HSF2的蛋白质通过与PP2A的PR65/A亚基结合来调节PP2A的活性，并通过直接与催化亚基竞争其在PR65中的结合位置来阻止其与催化亚单位的结合。结果表明，阻止催化亚基结合的Pr65突变与人类肺癌和结肠癌有关，证明了HSF2的S阻断Pr65与催化亚基相互作用的能力具有生物学意义。HSF2也是一个已知的转录因子，调节热休克蛋白(HSP)基因的表达，因此我们假设HSF2对PP2A的调控可能是HSP表达调控和细胞生长调控途径之间的一种串扰机制。为了阐明HSF2作为一种新的PP2A调节蛋白的功能，我们建议1)鉴定HSF2和PR65中对这种相互作用重要的序列，2)表征调节HSF2：PR65相互作用的细胞机制，3)确定HSF2在调节细胞PP2A活性中的功能，以及4)使用酵母双杂交筛选法(我们已经使用这种方法确定了一个新的候选蛋白，称为MEL-18)。这些研究将通过阐明PP2A调控的新机制以及细胞内两个重要信号通路-热休克蛋白(HSP)调控细胞生长/分化的新机制来促进知识的发展。这一知识有助于理解癌细胞的正常生长调控和异常增殖控制。

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.