Life Sciences Institute, Department of Biochemistry and Molecular Biology, The University of British Columbia, Vancouver, BC, Canada.
Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, VIC, Australia.
Commun Biol. 2021 Mar 5;4(1):291. doi: 10.1038/s42003-021-01830-x.
Pivotal to the maintenance of cellular homeostasis, macroautophagy (hereafter autophagy) is an evolutionarily conserved degradation system that involves sequestration of cytoplasmic material into the double-membrane autophagosome and targeting of this transport vesicle to the lysosome/late endosome for degradation. EPG5 is a large-sized metazoan protein proposed to serve as a tethering factor to enforce autophagosome-lysosome/late endosome fusion specificity, and its deficiency causes a severe multisystem disorder known as Vici syndrome. Here, we show that human EPG5 (hEPG5) adopts an extended "shepherd's staff" architecture. We find that hEPG5 binds preferentially to members of the GABARAP subfamily of human ATG8 proteins critical to autophagosome-lysosome fusion. The hEPG5-GABARAPs interaction, which is mediated by tandem LIR motifs that exhibit differential affinities, is required for hEPG5 recruitment to mitochondria during PINK1/Parkin-dependent mitophagy. Lastly, we find that the Vici syndrome mutation Gln336Arg does not affect the hEPG5's overall stability nor its ability to engage in interaction with the GABARAPs. Collectively, results from our studies reveal new insights into how hEPG5 recognizes mature autophagosome and establish a platform for examining the molecular effects of Vici syndrome disease mutations on hEPG5.
对细胞内稳态的维持至关重要，巨自噬（以下简称自噬）是一种进化上保守的降解系统，涉及细胞质物质被隔离到双层自噬体中，并将这种运输囊泡靶向溶酶体/晚期内体进行降解。EPG5 是一种大型后生动物蛋白，被认为是一种固定因子，以强制自噬体-溶酶体/晚期内体融合的特异性，其缺乏会导致一种严重的多系统疾病，称为 Vici 综合征。在这里，我们表明人类 EPG5（hEPG5）采用了扩展的“牧羊人手杖”结构。我们发现 hEPG5 优先结合对自噬体-溶酶体融合至关重要的人类 ATG8 蛋白 GABARAP 亚家族成员。hEPG5-GABARAPs 相互作用是由串联 LIR 基序介导的，这些基序具有不同的亲和力，对于 PINK1/Parkin 依赖性线粒体自噬中 hEPG5 向线粒体的募集是必需的。最后，我们发现 Vici 综合征突变 Gln336Arg 既不影响 hEPG5 的整体稳定性，也不影响其与 GABARAPs 相互作用的能力。总之，我们的研究结果揭示了 hEPG5 如何识别成熟自噬体的新见解，并为研究 Vici 综合征疾病突变对 hEPG5 的分子影响建立了一个平台。
