Proteasome-mediated degradation of misfolded proteins prevents aggregation inside and outside mitochondria. But how do cells safeguard the mitochondrial proteome and mitochondrial functions despite increased aggregation during proteasome inactivation? Here, using a novel two-dimensional complexome profiling strategy, we report increased supraorganization of respiratory complexes (RCs) in proteasome-inhibited cells that occurs simultaneously with increased pelletable aggregation of RC subunits inside mitochondria. Complex II (CII) and complex V (CV) subunits are increasingly incorporated into oligomers. Complex I (CI), complex III (CIII) and complex IV (CIV) subunits are engaged in supercomplex formation. We unravel unique quinary states of supercomplexes during early proteostatic stress that exhibit plasticity and inequivalence of constituent RCs. The core stoichiometry of CI and CIII is preserved, whereas the composition of CIV varies. These partially disintegrated supercomplexes remain functionally competent via conformational optimization. Subsequently, increased stepwise integration of RC subunits into holocomplexes and supercomplexes re-establishes steady-state stoichiometry. Overall, the mechanism of increased supraorganization of RCs mimics the cooperative unfolding and folding pathways for protein folding, but is restricted to RCs and is not observed for any other mitochondrial protein complexes.

This article has an associated First Person interview with the first author of the paper.

蛋白酶体介导的错误折叠蛋白的降解可防止线粒体内外聚集。但是，尽管在蛋白酶体失活过程中聚集增加，细胞如何保护线粒体蛋白质组和线粒体功能呢？在这里，使用一种新颖的二维复合物谱分析策略，我们报道了蛋白酶体抑制细胞中呼吸复合物（RCs）的超组织化增加，而线粒体内的RC亚基的可沉淀性聚集同时发生。复合物II（CII）和复合物V（CV）亚基越来越多地掺入低聚物中。复合物I（CI），复合物III（CIII）和复合物IV（CIV）亚基参与超复合物的形成。我们揭示了在早期蛋白水解应激过程中表现出可塑性和不等式RC的超复合物的独特五元态。CI和CIII的核心化学计量得以保留，而CIV的组成却有所不同。这些部分崩解的超复合物通过构象优化保持功能上的能力。随后，逐步增加的RC亚基逐步整合到全络合物和超络合物中，可重新建立稳态化学计量。总体而言，RCs超组织性增加的机制模拟了蛋白质折叠的协同展开和折叠途径，但仅限于RCs，其他线粒体蛋白复合物均未观察到。RC亚基逐步整合到全络合物和超络合物中的过程逐步增强，从而重新建立了稳态化学计量。总体而言，RCs超组织性增加的机制模拟了蛋白质折叠的协同展开和折叠途径，但仅限于RCs，其他线粒体蛋白复合物均未观察到。RC亚基逐步整合到全络合物和超络合物中的过程逐步增强，从而重新建立了稳态化学计量。总体而言，RCs超组织性增加的机制模拟了蛋白质折叠的协同展开和折叠途径，但仅限于RCs，其他线粒体蛋白复合物均未观察到。

本文与论文的第一作者进行了第一人称访谈。