Control of NF-AT Signaling Through Nuclear Import

描述(申请人提供)：核因子-AT转录因子是关键 T细胞激活的钙信号通路中的效应器， 肌肉发育和突触可塑性。人们普遍认为， 四种核因子-AT在静息细胞中是胞质的，但被触发进入细胞核 并通过钙调神经磷酸酶(一种钙激活的磷酸酶)反式激活基因。近期 然而，数据正在推翻NF-AT模型的一些假设，并 为管理这一部门的复杂监管机制提供了新的见解 钙信号。例如，小鼠对四种核因子-AT的基因敲除 同种异型得出了出人意料的结论，NFAT1和NF-AT4在 事实是T细胞激活的负调节因子，而只有核因子-AT2似乎能 对细胞因子基因具有预期的正活性。这些数据是 特别有趣的是，T细胞、心肌细胞和神经元似乎 同时表达多种核因子-AT亚型，表明激活必须 有赖于这些因素的不同调控。我们现在已经开始 通过评估所有四个NE-AT同种类型的动力学作为 钙信号与细胞环境的作用。值得注意的是，这些 数据揭示了NE-AT同型依赖的显著程度，并提示 细胞类型和钙非依赖途径在血管紧张素转换酶中的重要作用 最终的核因子-AT反应。因此，我们假设，这些核因子-AT家族 成员必须不同地受制于超越或 扩展那些由钙信号传递的信号，并理解这些 替代机制对于预测依赖于细胞功能是必不可少的 在核因子-ATS上。 这是一项揭示差异NE-AT信令机制的建议 牢房。我们将(1)，确定潜在的生化信号通路 NE-AT的差异调节及其程度；(2)阐明 通过高度保守的激活来运作的钙非依赖性通路 四种NE-AT的保守机制；(3)，通过 NE-AT动态差动调制的高通量光屏 NE-AT亚型；(4)利用这些小分子对其进行功能解剖 NE-AT亚型对T等复杂过程的个体贡献 细胞激活。我们预计，这项对NE-AT活性的新评估 将直接为自身免疫性疾病的更具体的治疗做出贡献， 器官移植和心肌病。

DESCRIPTION (provided by applicant): The NF-AT transcription factors are key effectors in the calcium signaling pathways underlying T cell activation, muscle development, and synaptic plasticity. It is generally thought that the four NF-ATs are cytoplasmic in resting cells but triggered to enter the nucleus and transactivate genes by calcineurin, a calciumactivated phosphatase. Recent data, however, are overturning some of the assumptions of the NF-AT model and offering new insights to the complex regulatory mechanisms governing this arm of calcium signaling. For instance, mouse gene knockouts of the four NF-AT isotypes have rendered the unexpected conclusion that NFAT1 and NF-AT4 are in fact negative regulators of T cell activation, while only NF-AT2 appears to have the predicted positive activity towards cytokine genes. These data are especially interesting because T cells, myocytes, and neurons appear to simultaneously express multiple NF-AT isotypes, indicating that activation must depend on the differential regulation of these factors. We have now begun to address this problem by evaluating the dynamics of all four NE-AT isotypes as a function of calcium signaling and cellular environment. Significantly, these data reveal a remarkable degree of NE-AT isotype-dependent dynamics and suggest important contributions of cell type and calcium-independent pathways to the ultimate NF-AT response. We hypothesize, therefore, that these NF-AT family members must be differentially subject to regulatory pathways that override or extend those imparted by calcium signaling, and that understanding these alternative mechanisms is essential for predicting cellular functions dependent on NF-ATs. This is a proposal to uncover the mechanisms of differential NE-AT signaling in the cell. We will (1), determine the biochemical signaling pathways underlying differential NE-AT regulation and their extent; (2) elucidate the calcium-independent pathways that operate through a highly conserved activation mechanism conserved in all four NE-ATs; (3), identify small molecules through high-throughput optical screens of NE-AT dynamics which differentially modulate the NE-AT isotypes; and (4), use these small molecules to functionally dissect the individual contributions of NE-AT isotypes to complex processes such as T cell activation. We anticipate that this novel evaluation of NE-AT activity will directly contribute to more specific therapies for autoimmune disease, organ transplantation, and cardiomyopathies.