Department of Biochemistry and Molecular Biology, Seoul National University College of Medicine, Seoul 03080, Republic of Korea; Department of Biomedical Sciences, Seoul National University Graduate School, Seoul 03080, Republic of Korea.
Department of Physiology and Biomedical Sciences, Seoul National University College of Medicine, Seoul 03080, Republic of Korea.
Biochim Biophys Acta Gene Regul Mech. 2023 Jun;1866(2):194932. doi: 10.1016/j.bbagrm.2023.194932. Epub 2023 Mar 28.
The spatiotemporal sequestration of misfolded proteins is a mechanism by which cells counterbalance proteome homeostasis upon exposure to various stress stimuli. Chronic inhibition of proteasomes results in a large, juxtanuclear, membrane-less inclusion, known as the aggresome. Although the molecular mechanisms driving its formation, clearance, and pathophysiological implications are continuously being uncovered, the biophysical aspects of aggresomes remain largely uncharacterized. Using fluorescence recovery after photobleaching and liquid droplet disruption assays, we found that the aggresomes are a homogeneously blended condensates with liquid-like properties similar to droplets formed via liquid-liquid phase separation. However, unlike fluidic liquid droplets, aggresomes have more viscosity and hydrogel-like characteristics. We also observed that the inhibition of aggresome formation using microtubule-disrupting agents resulted in less soluble and smaller cytoplasmic speckles, which was associated with marked cytotoxicity. Therefore, the aggresome appears to be cytoprotective and serves as a temporal reservoir for dysfunctional proteasomes and substrates that need to be degraded. Our results suggest that the aggresome assembles through distinct and potentially sequential processes of energy-dependent retrograde transportation and spontaneous condensation into a hydrogel.
错误折叠蛋白的时空隔离是细胞在暴露于各种应激刺激时平衡蛋白质组平衡的一种机制。蛋白酶体的慢性抑制会导致一个大的、核周的、无膜的包含物，称为聚集体。尽管驱动其形成、清除和病理生理影响的分子机制不断被揭示，但聚集体的生物物理方面在很大程度上仍未被描述。通过荧光恢复后光漂白和液滴破坏实验，我们发现聚集体是一种均匀混合的凝聚物，具有类似于通过液-液相分离形成的液滴的液体性质。然而，与流动的液滴不同，聚集体具有更高的粘度和水凝胶样的特性。我们还观察到，使用微管破坏剂抑制聚集体的形成会导致细胞质斑点的溶解度降低和体积减小，这与明显的细胞毒性有关。因此，聚集体似乎具有细胞保护作用，是功能失调的蛋白酶体和需要降解的底物的临时储存库。我们的结果表明，聚集体通过能量依赖的逆行运输和自发凝聚成水凝胶的独特且可能是连续的过程组装而成。
