Regulation of lymphocyte survival by NF-kB proteins

描述（由申请人提供）：淋巴细胞的发育和功能严重依赖于细胞凋亡对其存活的调节。过去几年开展的研究表明，TNF受体超家族成员（如TNF受体1和Fas）在活化的成熟淋巴细胞中起着凋亡的关键诱导剂的作用。然而，尚不清楚这些凋亡通路是否对发育中的淋巴细胞的存活也有重要的调节作用。本研究的目的是为了更好地了解参与淋巴细胞存活的促凋亡和抗凋亡机制。具体来说，我们将使用敲除小鼠模型研究凋亡调节NF-kB转录因子在调节发育中的b淋巴细胞存活中的作用。1)我们在这里展示了将ReIA-/-胎儿肝细胞过继转移到辐照淋巴细胞缺陷的Rag1-/-小鼠后的淋巴生成损伤。在缺乏Re1A和TNF受体1 （TNFR1）的情况下，受损的淋巴细胞生成在很大程度上得以恢复，这表明Re1A的一个关键功能可能是阻止TNFR1诱导的发育中的淋巴细胞的杀伤。在这里，我们将研究TNF诱导杀死RelA-/-发育中的B细胞的机制，包括半胱天冬酶蛋白酶和活性氧（ROS）在TNF杀伤中的潜在参与。2)为了了解RelA-/- B细胞对TNF易感性的机制，我们研究了可能重要的抗凋亡基因的调控。我们发现，在Re1A-/- B细胞中，一个关键的抗凋亡基因BcI-2的组成性表达显著降低。在这里，我们将进一步研究Re1A在调控潜在的重要抗凋亡基因中的可能作用。利用逆转录病毒转导系统，我们将在RelA-/- B细胞中重新表达表达受损的抗凋亡基因，以确定其对TNF杀伤的影响。逆转录病毒转导的RelA-/-胎肝造血前体在Rag1-/-小鼠中恢复淋巴细胞生成的能力也将被确定。3)我们还发现成熟T细胞在调节发育中的淋巴细胞存活方面具有潜在的重要作用。我们的研究结果表明，成熟的T细胞可能提供信号，可以抵消TNFR1的细胞毒性作用。在这里，我们将确定所涉及的T细胞亚群，以及负责这种保护作用的T细胞衍生因子的性质。作为这种T细胞保护作用的介质，CD4OL和IL-3特别有趣。在这里，我们将确定CD4OL和IL-3在体外抑制肿瘤坏死因子杀死Re1A-/-发育中的B细胞以及在体内挽救淋巴细胞生成中的作用。这些研究也可能为在治疗环境下增强淋巴系统生成的策略的发展提供见解。

DESCRIPTION (provided by applicant): The development and function of lymphocytes is critically dependent upon regulation of their survival by apoptosis. Studies 'carried-out over the last several years have shown that members of the TNF receptor superfamily (e.g., TNF receptor 1 and Fas) function as key inducers of apoptosis in activated mature lymphocytes. However, it is not known whether these apoptotic pathways 'are also important for regulating survival of developing lymphocytes. The goal of this investigation is to provide a better understanding of pro-apoptotic and anti-apoptotic mechanisms involved in developing lymphocyte survival. Specifically, we will study the role of the apoptosis-regulating NF-kB transcription factor in regulating survival of developing B-lymphocytes, using knock-out mouse models. 1) We show here an impairment of lymphopoiesis following adoptive transfer of ReIA-/- fetal liver cells into irradiated lymphocyte-deficient Rag1-/- mice. Impaired lymphocyte generation was largely rescued in the combined absence of Re1A and TNF receptor 1 (TNFR1), indicating a key function of Re1A may be to prevent TNFR1-induced killing of developing lymphocytes. Here we will investigate the mechanisms by which TNF induces killing of RelA-/- developing B cells, including the potential involvement of caspase proteases and reactive oxygen species (ROS) in TNF killing.2) To understand mechanisms responsible for susceptibility of RelA-/- B cells to TNF, we have studied regulation of potentially important anti-apoptotic genes. We have found that constitutive expression of one key anti-apoptotic gene, BcI-2, was 'significantly reduced in Re1A-/- B cells. Here we will further investigate a possible role for Re1A in regulation of potentially important anti-apoptotic genes. Using a retroviral transduction system, we will re-express anti-apoptotic genes showing impaired expression in RelA-/- B cells to determine their effect on TNF killing. The ability of retrovirus-transduced RelA-/- fetal liver hematopoietic precursors in restoring lymphocyte generation in Rag1-/- mice will also be determined.3) We have also identified a potentially important role for mature T cells in regulating survival of developing lymphocytes. Our results suggest that mature T cells may provide signals, which can counteract the cytotoxic effects of TNFR1. Here we will identify T cell subsets involved, and the nature of T cell derived factors responsible for this protective effect. Of particular interest as mediators of this T cell protective effect are CD4OL and IL-3. Here we will determine the role of CD4OL and IL-3 in inhibition of TNF killing of Re1A-/- developing B cells in vitro, and in rescuing Iymphopoiesis in vivo. These studies may also provide insights into development of strategies for enhancing lymphopoiesis under therapeutic settings.