Biochemical and Spatial Regulation of IKKg/NEMO During T Cell Activation

描述(由申请人提供)：需要对核因子-κB转录因子进行适当的调节，以启动对病原体的免疫反应，并保持免疫系统的正常内稳态。相反，过度依赖于NF-κB的炎症可能有助于推动自身免疫性疾病的发展和肿瘤的形成。在这种情况下，重要的是要知道NF-κB家族转录因子是如何通过影响这一途径的不同类型的受体的信号来控制的。在抗原激活T细胞的过程中，TCR和CD28以及许多信号蛋白被重组成不同的结构，同时经历了广为人知的翻译后修饰，如磷酸化和泛素化。虽然IKK复合体聚集在免疫突触中心富含TCR的区域，但受体信号是由接触外围的TCR微簇启动的。我们发现IKK复合体迅速被招募到含有TCR和ZAP70的信号微簇中，而不是SLP-76。我们的成像研究还揭示了更多的IKKγ池，这些池可能参与将IKKγ传递到TCR微簇和/或其下调。然而，关于IKKγ的亚细胞调控及其在T细胞激活中的作用，仍有许多重要的问题需要回答。如果我们最终要利用这一知识来操纵tcr介导的NF-κB激活用于治疗目的，进行这样的研究是很重要的。我们推测，经典的NF-κB通路的TcR/CD28激活需要复杂的亚细胞调控，即接头蛋白IKKγ/NEMO。因此，在目标1中，我们将确定ZAP70和LCK在将IKK复合体招募到TCR微簇中的作用。在目标2中，我们将确定Carma1及其相关蛋白在IKK复合体空间调控中的作用。最后，在目标3中，我们将定义包含总IKK复合体和活性IKK复合体的亚细胞隔室(S

DESCRIPTION (provided by applicant): Proper regulation of NF-κB transcription factors is required to mount immune responses to pathogens and for the normal homeostasis of the immune system. Conversely, excessive NF-κB-dependent inflammation can help to drive the development of autoimmune disease and the formation of tumors. In this light, it is important to know how NF-κB family transcription factors are controlled by signaling through the different types of receptors that impinge on this pathway. During T cell activation by antigen, TCR and CD28, and numerous signaling proteins, are reorganized into distinct structures while undergoing well-described post-translational modifications, such as phosphorylation and ubiquitination. Although the IKK complex accumulates in the TCR-rich domain at the center of the immune synapse, receptor signals are initiated by TCR 'microclusters' in the periphery of the contact. We have found that the IKK complex is rapidly recruited to signaling microclusters containing the TCR and Zap70, but not SLP-76. Our imaging studies have also revealed additional pools of IKKγ that may be involved in delivery of IKKγ to TCR microclusters and/or its down-regulation. However, a number of important questions remain to be answered regarding the sub-cellular regulation of IKKγ and its role in T cell activation. Carrying out such studies is important, if we are to eventually exploit this knowledge to manipulate TCR-mediated NF-κB activation for therapeutic purposes. We hypothesize that TCR/CD28 activation of the classical NF-κB pathway requires complex sub-cellular regulation of the adaptor protein IKKγ/NEMO. Thus, in Aim 1, we will determine the role of Zap70 and Lck in recruitment of the IKK complex to TCR microclusters. In Aim 2, we will define the role of Carma1 and associated proteins in spatial regulation of the IKK complex. Finally, in Aim 3 we will define the sub-cellular compartment(s) containing total and active IKK complexes