微丝细胞骨架调控凋亡细胞吞噬的机制研究

凋亡细胞清除对维持多细胞生物器官动态稳定有重要意义。在吞噬细胞内，两条保守信号通路共同激活小G蛋白CED-10，重排微丝细胞骨架，实现对凋亡细胞的吞噬。然而CED-10调控下游微丝骨架组装的机制尚不清楚。凋亡细胞的吞噬通常依赖于高度动态的分枝状微丝骨架，其组装由ARP2/3复合物和成核促进因子WASP和WAVE家族蛋白激活。本课题拟利用线虫凋亡Q细胞和吞噬细胞hyp 7为模型，研究CED-10调控微丝骨架重排机制。申请人发现CED-10通过与WASP结合调控其在凋亡Q细胞表面的富集；且WASP结合蛋白SEM-5通过促进分叉状微丝形成参与调控凋亡Q细胞清除。以此为基础，本课题将结合CRISPR-Cas9基因编辑技术，显微成像技术和生化分析，鉴定凋亡Q细胞清除过程中微丝调控因子，深入研究CED-10与这些因子的关系，拓展对微丝调控因子在凋亡细胞清除中的认识，为理解细胞凋亡相关疾病提供新思路。

The efficient removal of apoptotic cells by phagocytes is essential for tissue homeostasis in metazoans. Engulfment of apoptotic cells in Caenorhabditis elegans is controlled by two conserved genetic pathways, which activate CED-10/Rac to reorganize the actin cytoskeleton during phagocytosis; however, little is known about how CED-10/Rac is linked to actin reorganization. Phagocytosis requires highly dynamic branched actin filament networks, which are nucleated by the ARP2/3 (actin-related protein 2/3) complex and regulated by actin nucleation promoting factor WASP (Wiskott–Aldrich syndrome protein) and WAVE (WASP-family verprolin homologous protein) family proteins. The proposed study aims to elucidate the mechanism underlying actin reorganization when the C. elegans apoptotic Q cells are engulfed by the hypodermal hyp 7 cells. Our preliminary results have shown that WASP and ARP2/3 were recruited to apoptotic Q cells and that the clearance of apoptotic Q cells was delayed in WASP mutants. Our biochemical analysis has revealed that CED-10 interacted with WASP. In addition, we have uncovered that a WASP-associated protein SEM-5/GRB2 promoted the formation of branched F-actin in vitro and was required for the removal of apoptotic Q cell in vivo. We will combine the CRISPR-Cas9-based genome editing, fluorescence live imaging and biochemistry to identify actin polymerization regulators required for the clearance of apoptotic cells. We will further investigate how CED-10 regulates these actin regulators. We expect that our results will advance our understanding of actin regulators in corpse removal and provide new insights into the related diseases.