College of Pharmacy, Jinan University, Guangzhou, China.
Biochem Biophys Res Commun. 2013 Aug 2;437(3):482-8. doi: 10.1016/j.bbrc.2013.06.111. Epub 2013 Jul 9.
Autophagy plays a crucial role in a wide array of physiological processes. To uncover the complex regulatory networks and mechanisms underlying basal autophagy, we performed a quantitative proteomics analysis of autophagy-deficient mouse embryonic fibroblast cells (MEFs) using iTRAQ labeling coupled with on-line 2D LC/MS/MS. We quantified a total of 1234 proteins and identified 114 proteins that were significantly altered (90% confidence interval), including 48 up-regulated proteins and 66 down-regulated proteins. We determined that F-actin was disassembled in autophagy-deficient Atg7(-/-) MEFs. Treatment of the WT MEFs with cytochalasin D (CD), which induces F-actin depolymerization, significantly induced autophagosome formation. However, treatment with cytochalasin D also increased the protein level of p62 under starvation conditions, suggesting that depolymerization of F-actin impaired autophagosome maturation and that the intact F-actin network is required for basal and starvation-induced autophagy. Our results demonstrate a close relationship between F-actin and autophagy and provide the basis for further investigation of their interactions.
自噬在广泛的生理过程中起着至关重要的作用。为了揭示基础自噬的复杂调控网络和机制，我们使用 iTRAQ 标记结合在线 2D LC/MS/MS 对自噬缺陷型小鼠胚胎成纤维细胞（MEFs）进行了定量蛋白质组学分析。我们总共定量了 1234 种蛋白质，鉴定出 114 种显著改变的蛋白质（90%置信区间），包括 48 种上调的蛋白质和 66 种下调的蛋白质。我们发现自噬缺陷型 Atg7(-/-) MEFs 中的 F-肌动蛋白解聚。用细胞松弛素 D（CD）处理 WT MEFs，诱导 F-肌动蛋白解聚，可显著诱导自噬体形成。然而，在用细胞松弛素 D 处理饥饿条件下时，p62 的蛋白水平也增加，表明 F-肌动蛋白的解聚会损害自噬体的成熟，并且完整的 F-肌动蛋白网络是基础自噬和饥饿诱导自噬所必需的。我们的结果表明 F-肌动蛋白和自噬之间存在密切关系，并为进一步研究它们的相互作用提供了基础。
