張宏

摘 要：該研究以秀麗線蟲(chóng)為模型，以細(xì)胞自吞噬為中心，用遺傳學(xué)、細(xì)胞生物學(xué)、生化、蛋白質(zhì)組學(xué)等多種手段全面系統(tǒng)地鑒定多細(xì)胞生物中參與自吞噬過(guò)程的所有組成基因，研究自吞噬的調(diào)節(jié)機(jī)制。同時(shí)，還著重研究自吞噬與另外兩個(gè)基本的生命過(guò)程——細(xì)胞凋亡和個(gè)體衰老之間的相互關(guān)系和調(diào)節(jié)作用。該研究共分為4部分：第一，建立了以秀麗線蟲(chóng)為研究細(xì)胞自噬的多細(xì)胞生物模型，通過(guò)遺傳學(xué)手段篩選并克隆新的細(xì)胞自噬分子，闡明其作用的分子、細(xì)胞及遺傳調(diào)控機(jī)制，揭示這些新分子在個(gè)體發(fā)育過(guò)程中的功能，并為哺乳動(dòng)物細(xì)胞中的同源基因的研究提供線索。第二，研究了凋亡細(xì)胞清除的多個(gè)環(huán)節(jié)，發(fā)現(xiàn)了在凋亡細(xì)胞清除的不同階段發(fā)揮作用的新調(diào)控因子并闡明了其作用機(jī)制。成果包括：橋聯(lián)分子TTR-52介導(dǎo)凋亡細(xì)胞識(shí)別及吞噬的調(diào)控機(jī)制；TTR-52、CED-7及脂結(jié)合/轉(zhuǎn)運(yùn)蛋白NRF-5協(xié)同調(diào)控PS，然后吞噬細(xì)胞表面的呈現(xiàn)；肌管素磷酸酶MTM-1負(fù)調(diào)控凋亡細(xì)胞吞噬。第三，通過(guò)比較長(zhǎng)壽daf-2突變體線蟲(chóng)和野生型線蟲(chóng)的線粒體，包括形態(tài)、生理活性特征、線粒體DNA的拷貝數(shù)和線粒體蛋白質(zhì)組的差異，找出daf-2突變體長(zhǎng)壽的原因，探索在多細(xì)胞動(dòng)物中細(xì)胞自體吞噬與衰老之間的關(guān)系，以及胰島素信號(hào)調(diào)節(jié)線粒體和細(xì)胞自吞噬的作用機(jī)制。第四，開(kāi)發(fā)和完善了各項(xiàng)基于生物質(zhì)譜的蛋白質(zhì)組學(xué)技術(shù)，利用這些生物質(zhì)譜技術(shù)對(duì)哺乳動(dòng)物細(xì)胞自吞噬的起始進(jìn)行了深入的研究和探討。同時(shí)，還將開(kāi)發(fā)的生物質(zhì)譜技術(shù)廣泛應(yīng)用到各個(gè)重要的生物學(xué)領(lǐng)域中，推動(dòng)了生物學(xué)研究領(lǐng)域的發(fā)展。

關(guān)鍵詞：秀麗線蟲(chóng) 細(xì)胞自噬 細(xì)胞凋亡 衰老 質(zhì)譜

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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