Role of Bai1 and Elmo Proteins in Apoptotic Cell Clearance

描述(由申请者提供)：我们每天在体内更新数十亿个细胞，作为正常内稳态的一部分，其中许多细胞在凋亡过程中死亡。体内的吞噬细胞以一种“免疫沉默”的方式迅速清除了凋亡细胞。迅速去除的缺陷会导致继发性坏死，死亡细胞释放细胞内内容物促进慢性组织炎症，这与自身免疫性疾病有关，如系统性红斑狼疮、呼吸道炎症和动脉粥样硬化。因此，更好地了解凋亡细胞识别和清除的机制对于对抗这些疾病是必要的。目前的竞争性续签申请是根据前两个供资期间取得的进展提出的，每个供资期限为4年。我们首先鉴定了一种新的细胞质吞噬适配蛋白ELMO1，并将Elmo/Dock180/Rac信号通路定义为促进细胞凋亡吞噬的进化保守模块。然后，我们确定膜蛋白BAI1是ELMO1上游的吞噬受体；独立地，我们为ELMO1产生了敲除小鼠，以确定它在睾丸中的凋亡生殖细胞清除和大脑中的神经发生过程中的需求。这些工作还提出了一些令人兴奋的下一组问题，涉及体内通过BAI1/ELMO1模块的信号传递，以及这一途径如何抑制组织中的炎症。在这项建议的Aim1中，使用我们实验室最近产生的BAI1基因被靶向缺失(花蕾和直接敲除)的小鼠，我们将测试BAI1在体内两个不同组织--胸腺和睾丸--的凋亡细胞清除中发挥作用的假设。此外，使用过度表达野生型或突变形式的BAI1的可诱导转基因小鼠，我们将测试BAI1在促进体内吞噬方面是否提供独特的与冗余的信号。目的2重点研究细胞凋亡识别如何转化为抗炎细胞因子的产生，这是细胞凋亡清除的关键特征，目前还不完全清楚。我们将验证BAI1-ELMO1介导的信号有助于吞噬细胞抗炎反应的假设。具体地说，我们将测试ELMO1和转录机制组件(我们在初步研究中发现)之间的意外相互作用如何有助于吞噬过程中的抗炎基因转录。我们将在体内使用Elmo2缺陷的小鼠(我们最近培育的)在组织炎症的小鼠模型中推广这些方法。总而言之，我们期望这些研究能在体内细胞清除中通过BAI1/ELMO1信号通路提供令人兴奋的新信息。由于ELMO1和BAI1的表达改变与人类的炎症性疾病有遗传联系，因此拟议的研究结果可能与炎症性疾病的治疗干预相关。

DESCRIPTION (provided by applicant): We turnover billions of cells in the body each day as part of normal homeostasis, and many of these cells die by the process of apoptosis. The apoptotic cells are quickly cleared by phagocytes in vivo, in an 'immunologically silent' fashion. Defects in prompt removal leads to secondary necrosis, with the release of intracellular contents from the dying cells promoting chronic tissue inflammation, which has been linked to autoimmune disorders such as SLE, airway inflammation, and atherosclerosis. Thus, a better knowledge of the mechanisms of apoptotic cell recognition and clearance becomes necessary for countering these disorders. This current competitive renewal application is based on the progress made during the two previous funding periods of 4 years each. We first identified a novel cytoplasmic engulfment adapter protein ELMO1, and defined the ELMO/Dock180/Rac signaling pathway as an evolutionarily conserved module for promoting apoptotic cell engulfment. We then identified the membrane protein BAI1 as the engulfment receptor upstream of ELMO1; independently, we generated knockout mice for Elmo1 to identify its requirement in apoptotic germ cell clearance in the testes, and during neurogenesis in the brain. These works have also raised a number of exciting next set of questions on signaling via the BAI1/ELMO1 module in vivo, and how this pathway may dampen inflammation in tissues. In Aim1 of this proposal, using mice recently generated in our laboratory where the Bai1 locus has been targeted for deletion (floxed as well as straight knockout), we will test the hypothesis that BAI1 plays a role in apoptotic cell clearance in vivo in two different tissues, the thymus and the testes. Moreover, using inducible transgenic mice that overexpress wild type or a mutant form of BAI1, we will test whether BAI1 provides unique versus redundant signals in promoting engulfment in vivo. Aim 2 focuses on how apoptotic cell recognition translates to anti-inflammatory cytokine production, key feature of apoptotic cell clearance that is not fully understood. We will test the hypothesis that the BAI1-ELMO1-mediated signaling contributes to anti-inflammatory responses of phagocytes. Specifically, we will test how an unexpected interaction between ELMO1 and components of the transcriptional machinery (which we have discovered in our preliminary studies), contribute to the anti-inflammatory gene transcription during engulfment. We will extend these in vivo using mice deficient in Elmo2 (that we have recently generated) in a mouse model of tissue inflammation. Collectively, we expect these studies to provide exciting new information on signaling via the BAI1/Elmo1 signaling pathway in cell clearance in vivo. Since altered expression of ELMO1 and BAI1 are genetically linked to inflammatory disorders in humans, the results from the proposed studies could be relevant for therapeutic intervention in inflammatory disorders.