多细胞生物吞噬的调节机制

张宏

摘 要：该研究以秀丽线虫为模型，以细胞自吞噬为中心，用遗传学、细胞生物学、生化、蛋白质组学等多种手段全面系统地鉴定多细胞生物中参与自吞噬过程的所有组成基因，研究自吞噬的调节机制。同时，还着重研究自吞噬与另外两个基本的生命过程——细胞凋亡和个体衰老之间的相互关系和调节作用。该研究共分为4部分：第一，建立了以秀丽线虫为研究细胞自噬的多细胞生物模型，通过遗传学手段筛选并克隆新的细胞自噬分子，阐明其作用的分子、细胞及遗传调控机制，揭示这些新分子在个体发育过程中的功能，并为哺乳动物细胞中的同源基因的研究提供线索。第二，研究了凋亡细胞清除的多个环节，发现了在凋亡细胞清除的不同阶段发挥作用的新调控因子并阐明了其作用机制。成果包括：桥联分子TTR-52介导凋亡细胞识别及吞噬的调控机制；TTR-52、CED-7及脂结合/转运蛋白NRF-5协同调控PS，然后吞噬细胞表面的呈现；肌管素磷酸酶MTM-1负调控凋亡细胞吞噬。第三，通过比较长寿daf-2突变体线虫和野生型线虫的线粒体，包括形态、生理活性特征、线粒体DNA的拷贝数和线粒体蛋白质组的差异，找出daf-2突变体长寿的原因，探索在多细胞动物中细胞自体吞噬与衰老之间的关系，以及胰岛素信号调节线粒体和细胞自吞噬的作用机制。第四，开发和完善了各项基于生物质谱的蛋白质组学技术，利用这些生物质谱技术对哺乳动物细胞自吞噬的起始进行了深入的研究和探讨。同时，还将开发的生物质谱技术广泛应用到各个重要的生物学领域中，推动了生物学研究领域的发展。

关键词：秀丽线虫 细胞自噬 细胞凋亡 衰老 质谱

Mechanism of Autophagy in Multicellular Organism

Zhang Hong

（National Institute of Biological Sciences， Beijing）

Abstract：We used C. elegans as a multicellular genetic model system to investigate molecular mechanisms of autophagy， apoptosis and aging. The project aimed to address the following questions： the molecular machinery of the evolutionarily conserved autophagy-lysosome pathway， the phagocytosis pathway and also how these processes affect the aging process during animal development. The research includes the following aspects： First， we established C. elegans as a multicellular genetic model to delineate the autophagic machinery by demonstrating that a variety of protein aggregates are selectively removed by autophagy during embryogenesis. We also investigated the physiological function of these metazoan-specific autophagy genes in mice. Second， we studied the phagocytic removal of apoptotic cells， which is an integral part of the cell death program and an important event in tissue remodeling， suppression of inflammation and regulation of the immune response. We have identified six novel regulators and revealed mechanisms through which they regulate various aspects of cell corpse removal. Among these newly identified regulators， TTR-52 and NRF-5 are extracellular lipid-binding/transfer proteins that mediate recognition of cell corpses. Third， we investigated the role of mitochondria in aging process. Using metabolic labeling with the stable heavy isotope 15N and quantitative mass spectrometry （MS）， we find that proteins up-regulated in daf-2 are highly enriched for those functioning in fatty acid metabolism， glyoxylate cycle， amino acid metabolism， or ROS metabolism. Fourth， we set up a high resolution biological mass spectrometer. According to the research directions proposed by the grant， we developed a variety of mass spectrometry based proteomics techniques. Through biological mass spectrometry， we identified multiple phosphorylation sites on proteins that help initiate autophagy in mammalian cells. Besides the field of autophagy， we also applied the biological mass spectrometric techniques in a variety of important biological fields including necrosis and aging， contributing greatly to the advancement of these fields. Our studies help us to understand the molecular autophagic pathway and also the physiological function in multicellular organisms and also how the autophagy-lysosomal pathway interacts with the endosomal-lysosomal pathway.

Key Words：C. elegans； Autophagy； Apoptosis； Aging； Mass

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