Super-resolution Microscopy Reveals Compartmentalization of Peroxisomal Membrane Proteins.
Super-resolution Microscopy Reveals Compartmentalization of Peroxisomal Membrane Proteins.
复制标题
超分辨率显微镜揭示了过氧化物酶体膜蛋白的分室化。
DOI：
10.1074/jbc.m116.734038
复制
发表时间：
2016-08-12
期刊：
影响因子：
--
通讯作者：
Eggeling C
中科院分区：
文献类型：
--
作者：
Galiani S;Waithe D;Reglinski K;Cruz-Zaragoza LD;Garcia E;Clausen MP;Schliebs W;Erdmann R;Eggeling C
过氧化物酶体蛋白输入过程中的膜相关事件在过氧化物酶体功能中起着至关重要的作用。由于缺少能够直接在细胞中研究过氧化物酶体的技术的空间分辨率，这些过程的许多细节尚不清楚。在这里，我们提出了使用超分辨率光学受激发射损耗显微镜调查与子60纳米分辨率的过氧化物酶体蛋白PEX5，PEX14，和PEX11周围积极进口过氧化物酶体的异质空间组织，这些过氧化物酶之间表现出明显的差异。此外，进口的蛋白固醇载体蛋白2（SCP 2）只占据了一个亚区的较大的过氧化物酶体，突出了蛋白质的异质性分布，甚至在过氧化物酶体。最后，我们的数据显示过氧化物酶体的亚群仅显示PEX14和PEX5或PEX11之间的弱共定位，但同时具有明显的区室化组织。这种区室化在强共定位的情况下不太明显，表明与过氧化物酶体中发生的变化有关的动态蛋白质重组。通过使用受激发射损耗显微镜，我们已经能够表征过氧化物酶体及其组分的细节，同时保持高度的统计方法，为将来同样复杂的生物学研究铺平了道路。
Membrane-associated events during peroxisomal protein import processes play an essential role in peroxisome functionality. Many details of these processes are not known due to missing spatial resolution of technologies capable of investigating peroxisomes directly in the cell. Here, we present the use of super-resolution optical stimulated emission depletion microscopy to investigate with sub-60-nm resolution the heterogeneous spatial organization of the peroxisomal proteins PEX5, PEX14, and PEX11 around actively importing peroxisomes, showing distinct differences between these peroxins. Moreover, imported protein sterol carrier protein 2 (SCP2) occupies only a subregion of larger peroxisomes, highlighting the heterogeneous distribution of proteins even within the peroxisome. Finally, our data reveal subpopulations of peroxisomes showing only weak colocalization between PEX14 and PEX5 or PEX11 but at the same time a clear compartmentalized organization. This compartmentalization, which was less evident in cases of strong colocalization, indicates dynamic protein reorganization linked to changes occurring in the peroxisomes. Through the use of multicolor stimulated emission depletion microscopy, we have been able to characterize peroxisomes and their constituents to a yet unseen level of detail while maintaining a highly statistical approach, paving the way for equally complex biological studies in the future.