Garin J, Diez R, Kieffer S, Dermine J F, Duclos S, Gagnon E, Sadoul R, Rondeau C, Desjardins M
Laboratoire de Chimie des protéines, Commissariat a l'Energie Atomique, 38054 Grenoble, France.
J Cell Biol. 2001 Jan 8;152(1):165-80. doi: 10.1083/jcb.152.1.165.
Phagosomes are key organelles for the innate ability of macrophages to participate in tissue remodeling, clear apoptotic cells, and restrict the spread of intracellular pathogens. To understand the functions of phagosomes, we initiated the systematic identification of their proteins. Using a proteomic approach, we identified >140 proteins associated with latex bead-containing phagosomes. Among these were hydrolases, proton pump ATPase subunits, and proteins of the fusion machinery, validating our approach. A series of unexpected proteins not previously described along the endocytic/phagocytic pathways were also identified, including the apoptotic proteins galectin3, Alix, and TRAIL, the anti-apoptotic protein 14-3-3, the lipid raft-enriched flotillin-1, the anti-microbial molecule lactadherin, and the small GTPase rab14. In addition, 24 spots from which the peptide masses could not be matched to entries in any database potentially represent new phagosomal proteins. The elaboration of a two-dimensional gel database of >160 identified spots allowed us to analyze how phagosome composition is modulated during phagolysosome biogenesis. Remarkably, during this process, hydrolases are not delivered in bulk to phagosomes, but are instead acquired sequentially. The systematic characterization of phagosome proteins provided new insights into phagosome functions and the protein or groups of proteins involved in and regulating these functions.
吞噬体是巨噬细胞参与组织重塑、清除凋亡细胞以及限制细胞内病原体传播的先天能力的关键细胞器。为了了解吞噬体的功能，我们开始对其蛋白质进行系统鉴定。采用蛋白质组学方法，我们鉴定出了140多种与含乳胶珠吞噬体相关的蛋白质。其中包括水解酶、质子泵ATP酶亚基以及融合机制的蛋白质，验证了我们的方法。我们还鉴定出了一系列先前未在内吞/吞噬途径中描述过的意外蛋白质，包括凋亡蛋白半乳糖凝集素3、Alix和肿瘤坏死因子相关凋亡诱导配体（TRAIL）、抗凋亡蛋白14-3-3、富含脂筏的flotillin-1、抗微生物分子乳粘连蛋白以及小GTP酶rab14。此外，有24个斑点的肽质量无法与任何数据库中的条目匹配，它们可能代表新的吞噬体蛋白质。构建一个包含160多个已鉴定斑点的二维凝胶数据库，使我们能够分析在吞噬溶酶体生物发生过程中吞噬体组成是如何被调节的。值得注意的是，在此过程中，水解酶并非大量输送到吞噬体，而是依次获取的。对吞噬体蛋白质的系统表征为吞噬体功能以及参与和调节这些功能的蛋白质或蛋白质组提供了新的见解。
