NOVEL CDK2 MECHANISM CONTROLS CELL PROLIFERATION

非转化T细胞需要IL-2来增殖，而il - 2rα是高亲和力生物相关IL-2R的必要成分。大量研究表明，在有丝分裂刺激的T细胞中，il - 2rα的转录上调。另一方面，il - 2rα表达的转录后控制尚未得到充分探索或明确证明，这具有潜在的生物学重要性。本研究提供的数据清楚地表明，IL-2Ralpha的表达在有丝分裂刺激下的原代T细胞的翻译水平上受到调节。我们发现il - 2rα转录物的翻译增加与IL-2信号通路的诱导是平行的，并且我们认为il - 2rα的翻译上调需要细胞周期调节激酶cdk2的活性。有趣的是，我们发现cdk2的表达也在转录后水平上受到控制，并且很可能是由cdk2依赖机制控制的。这些研究发现了cdk2的新作用，并表明以前未被认识的机制有助于IL-2Ralpha和cdk2的表达。我们还提供的数据显示cdk2激活和IL-2信号是脾T细胞中相互依赖的过程。我们认为cdk2活性对于IL-2R的有效翻译是必需的，并且IL-2R产生的信号通过维持cdk2抑制剂P27kip1的下调来促进cdk2的激活。这些相互作用和cdk2参与两个重要的细胞周期调节分子- IL-2Ralpha和cdk2本身-的翻译形成了我们的建议的重点。具体来说，我们将研究cdk2活性可能控制原代脾细胞中il - 2rα翻译的潜在机制。我们将确定il - 2rα mRNA的5'非翻译区负责翻译调节的区域，并将确定蛋白质是否以依赖于cdk2活性的方式与该区域结合。我们还将确定cdk2活性是否调节cdk2的合成或稳定性。进一步的实验将评估IL-2信号传导对tcr活化脾细胞中p27kip1持续下调的贡献。最后，TCR激活促进细胞周期蛋白A表达的机制将被描述。

Nontransformed T cells require IL-2 for proliferation, and IL-2Ralpha is an obligatory component of the high affinity biologically relevant IL-2R. Numerous studies have characterized the transcriptional up-regulation of IL-2Ralpha in mitogenically stimulated T cells. On the other hand, and of potential biological importance, post transcriptional control of IL-2Ralpha expression has yet to be adequately explored or definitively demonstrated. Data presented in this proposal clearly show that IL-2Ralpha expression is regulated at the translational level in primary T cells exposed to mitogenic stimuli. We show that increased translation of the IL-2Ralpha transcript is paralleled by the induction of IL-2 signaling pathways, and we suggest that translational up-regulation of IL-2Ralpha requires the activity of the cell cycle regulatory kinase, cdk2. Interestingly, we found that cdk2 expression was also controlled at a post- transcriptional level and, most likely, by a cdk2-dependent mechanism. These studies identify novel actions of cdk2 and suggest that previously unrecognized mechanisms contribute to the expression of IL-2Ralpha and cdk2. We also present data showing that cdk2 activation and IL-2 signaling are interdependent processes in splenic T cells. We suggest that cdk2 activity is required for the efficient translation of IL-2R and that signals generated by the IL-2R facilitate cdk2 activation by maintaining the down-regulation of the cdk2 inhibitor, P27kip1. These interactions and the involvement of cdk2 in the translation of two important cell cycle regulatory molecules - IL-2Ralpha and cdk2 itself - form the focus of our proposal. Specifically, we will examine potential mechanisms by which cdk2 activity might control the translation of IL-2Ralpha in primary splenocytes. We will identify the region of the 5' untranslated region of the IL-2Ralpha mRNA that is responsible for translational regulation and will determine if proteins bind to this region in a manner dependent on cdk2 activity. We will also determine if cdk2 activity modulates the synthesis or the stability of cdk2. Additional experiments will assess the contribution of IL-2 signaling to the continued down-regulation of p27kip1 in TCR-activated splenocytes. Lastly, the mechanism by which TCR activation promotes the expression of cyclin A will be delineated.