NF-KB REGULATION OF LYMPHOCYTE FUNCTION AND APOPTOSIS

在脊椎动物中，除了激活必需的先天防御机制外，NF-κ B/Rel因子被认为是适应性免疫的中枢协调调节因子。然而，由于先天免疫系统细胞中NF-κ B的伴随缺陷和NF-κ B复合物的冗余功能，不能通过分析携带单个NF-κ B基因破坏的小鼠来直接评估淋巴细胞中NF-κ B执行的广谱关键功能。 我们已经开发了一个模型系统，其中，由于高度同源的亚基p50和p52的同时丢失，NF-κ B二聚体的库被深深地扰乱。 我们建议使用该模型直接和系统地评价NF-κ B因子在B和T淋巴细胞发育和功能中的生物学影响。 该提案有三个具体目标。 在目标1中，我们将讨论NF-κ B在B淋巴细胞生成中的作用。 NF-κ B复合物是建立长寿命外周B细胞库所必需的。通过结合体内和体外方法，我们将确定NF-κ B控制该成熟步骤的机制，并将从评价先前提出的NF-κ B在B细胞存活、活化和有丝分裂中的作用的生理相关性开始。确定转录因子对B细胞选择的贡献也可以提供对自身免疫机制和治疗的见解。 在目标2中，我们将研究NF-κ B在T细胞稳态中的作用。 先前提出的NF-κ B在共刺激和活化诱导的细胞死亡（AICD）中的作用的生理相关性将通过检查NF-κ B缺陷型T细胞在受到完整的先天免疫系统指示时对其天然抗原的应答来确定。 为了进一步评估NF-κ B在自身免疫和慢性炎症中的潜在广泛作用，还将检查这些细胞的自身抗原驱动的应答。 在目标3中，我们试图确定和表征一个子集的NF-κ B调节基因的作用，以拮抗细胞凋亡。 为了实现这一目标，我们已经成功地优化了哺乳动物库的功能筛选的实验方案。 这是令人感兴趣的，因为这些基因似乎是由B和T细胞中的共刺激途径激活的存活程序的整体效应物，并且可能直接参与成熟B细胞和T细胞稳态的产生。 此外，NF-κ B调节的促存活基因与肿瘤发生、肿瘤细胞对化疗和电离辐射的抗性有关，从而为癌症治疗提供了潜在的新靶点。

In vertebrates, in addition to activating essential innate defense mechanisms, NF-kappaB/Rel factors are believed to function as central coordinating regulators of adaptive immunity. However, the direct assessment of the broad spectrum of critical functions executed by NF-kappaB in lymphocytes could not be obtained by the analyses of mice harboring disruptions of individual NF-kappaB genes, because of the concomitant deficiency of NF-kappaB in cells of the innate immune system and the redundant functions of NF-kappaB complexes. We have developed a model system where, due to the simultaneous loss of highly homologous subunits p50 and p52, the repertoire of NF-kappaB dimers is profoundly perturbed. We propose to use this model in a direct and systematic evaluation of the biologic impact of NF- kappaB factors in the development and function of B and T lymphocytes. The proposal has three Specific Aims. In Aim 1, we will address the role of NF-kappaB in B lymphopoiesis. NF-kappaB complexes are required for the establishment of the long-lived peripheral B cell pool By combining an in vivo and an in vitro approach, we will determine the mechanism(s) through which NF- kappaB controls this maturation step, and will begin by evaluating the physiologic relevance of previously proposed roles of NF-kappaB in B cell survival, activation, and mitogenesis. Defining the contribution of the transcript factor to B cell selection may also offer insights into autoimmune mechanisms and treatments. In Aim 2, we will examine roles of NF-kappaB in homeostasis of T cells. The physiologic relevance of previously suggested roles of NF-kappaB in costimulation and activation- induced cell death (AICD) will be determined by examining responses of NF-kappaB deficient T cells to their natural antigen, when instructed by an intact innate immune system. To further assess potential broad roles of NF-kappaB in autoimmunity and chronic inflammation, self-antigen-driven responses of these cells will also be examined. In Aim 3, we seek to identify and characterize a subset of NF-kappaB regulated genes that act to antagonize apoptosis. To achieve this goal we have successfully optimized an experimental protocol for the functional screening of mammalian libraries. This is of interest since these genes appear to be integral effectors of survival programs activated by costimulatory pathways in B and T cells, and may be directly involved in the generation of mature B cells and T cell homeostasis. In addition, NF-kappaB regulated pro-survival genes have been implicated in tumorigenesis, and resistance of tumor cells to chemotherapy and ionizing radiation, thereby providing potential new targets for cancer therapy.