Higher-order assemblies of oligomeric cargo receptor complexes form the membrane scaffold of the Cvt vesicle.
Higher-order assemblies of oligomeric cargo receptor complexes form the membrane scaffold of the Cvt vesicle.
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DOI：
10.15252/embr.201541960
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发表时间：
2016-07
期刊：
影响因子：
7.7
通讯作者：
Sachse C
中科院分区：
文献类型：
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作者：
Bertipaglia C;Schneider S;Jakobi AJ;Tarafder AK;Bykov YS;Picco A;Kukulski W;Kosinski J;Hagen WJ;Ravichandran AC;Wilmanns M;Kaksonen M;Briggs JA;Sachse C
选择性自噬是一种机制，通过这种机制，大的货物被专门隔离降解。货物和受体组装产生自噬囊泡的结构细节仍有待阐明。我们利用酵母细胞质到液泡靶向（Cvt）途径，选择性自噬的原型，以及多尺度分析方法来研究Cvt囊泡的分子结构。我们报告了主要Cvt货物Ape 1的寡聚性质，其具有2.8 μ m X射线和负染色EM结构，以及具有6.3 μ m cryo-EM结构的次要货物Ams 1。我们发现，主要的十二聚体货物prApe 1表现出形成高阶链结构的趋势，这些结构在体外与受体Atg 19相互作用时被破坏。这些货物-受体复合物的化学计量是维持体内Cvt聚集体大小的关键。使用相关的光学和电子显微镜，我们进一步可视化的CVT囊泡生物发生的关键阶段。我们的研究结果表明，Atg 19的相互作用限制了Ape 1聚集体的大小，同时作为空泡传递四聚体Ams 1的载体。
Selective autophagy is the mechanism by which large cargos are specifically sequestered for degradation. The structural details of cargo and receptor assembly giving rise to autophagic vesicles remain to be elucidated. We utilize the yeast cytoplasm‐to‐vacuole targeting (Cvt) pathway, a prototype of selective autophagy, together with a multi‐scale analysis approach to study the molecular structure of Cvt vesicles. We report the oligomeric nature of the major Cvt cargo Ape1 with a combined 2.8 Å X‐ray and negative stain EM structure, as well as the secondary cargo Ams1 with a 6.3 Å cryo‐EM structure. We show that the major dodecameric cargo prApe1 exhibits a tendency to form higher‐order chain structures that are broken upon interaction with the receptor Atg19 in vitro. The stoichiometry of these cargo–receptor complexes is key to maintaining the size of the Cvt aggregate in vivo. Using correlative light and electron microscopy, we further visualize key stages of Cvt vesicle biogenesis. Our findings suggest that Atg19 interaction limits Ape1 aggregate size while serving as a vehicle for vacuolar delivery of tetrameric Ams1.