Correlated cryogenic fluorescence microscopy and electron cryo-tomography shows that exogenous TRIM5α can form hexagonal lattices or autophagy aggregates in vivo.
Correlated cryogenic fluorescence microscopy and electron cryo-tomography shows that exogenous TRIM5α can form hexagonal lattices or autophagy aggregates in vivo.
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DOI：
10.1073/pnas.1920323117
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发表时间：
2020-11-24
影响因子：
11.1
通讯作者：
Jensen GJ
中科院分区：
文献类型：
--
作者：
Carter SD;Mamede JI;Hope TJ;Jensen GJ
TRIM5蛋白最显著的特征之一是其通过结合病毒衣壳来限制逆转录病毒感染的能力。TRIM5α会形成各种大小的高度动态的点状结构，并且在纯化时会在HIV病毒粒子表面形成六边形网，但细胞体的分子超微结构以及体外形成的网与HIV限制之间的关系仍不清楚。为了确定YFP - rhTRIM5α体的点状和动态性质背后的细胞超微结构，我们将低温相关光学和电子显微镜与电子冷冻断层扫描技术应用于TRIM5α体，并观察到YFP - rhTRIM5α定位在自噬途径的聚集体自噬分支中的细胞器上。与先前的研究一致，我们还发现TRIM5α在细胞内形成六边形网。
三联基序（TRIM）蛋白家族的成员已被证明可在细胞核和细胞质中组装成结构。TRIM蛋白家族的一个成员TRIM5α已被证明可形成参与限制诸如HIV - 1等逆转录病毒的细胞质体。在这里，我们将低温相关光学和电子显微镜与电子冷冻断层扫描技术应用于表达YFP - rhTRIM5α的完整哺乳动物细胞，发现存在六边形网，其臂长与纯化的TRIM5α在体外形成的六边形网的臂长相似。我们还在多种结构中观察到YFP - rhTRIM5α，这些结构具有参与巨自噬不同阶段的细胞器所预期的特征，包括无序的蛋白质聚集体（隔离体）、两侧有扁平双膜囊泡的隔离体（隔离体：吞噬泡复合物）、双膜囊泡内的隔离体（自噬体）以及多泡自噬泡内的隔离体（两性体或自溶酶体）。在这些结构中还看到了穹窿体，这与其在自噬中的作用一致。我们的数据1）支持最近的报道，即TRIM5α既可以形成组织良好的信号复合物，也可以形成非信号聚集体；2）提供了接近天然状态的巨自噬途径的图像；3）揭示了穹窿体在巨自噬早期就出现。
One of the most notable features of TRIM5 proteins is their ability to restrict retroviral infections by binding viral capsids. TRIM5α forms highly dynamic puncta of various sizes, and, when purified, hexagonal nets on the surface of HIV virions, but the molecular ultrastructure of the cellular bodies and the relationship of the in vitro nets to HIV restriction has remained unclear. To define the cellular ultrastructure underlying the punctate and dynamic nature of YFP-rhTRIM5α bodies, we applied cryogenic correlated light and electron microscopy combined with electron cryo-tomography to TRIM5α bodies and observed YFP-rhTRIM5α-localization to organelles found along the aggrephagy branch of the autophagy pathway. Consistent with previous work, we also found that TRIM5α forms hexagonal nets inside cells. Members of the tripartite motif (TRIM) protein family have been shown to assemble into structures in both the nucleus and cytoplasm. One TRIM protein family member, TRIM5α, has been shown to form cytoplasmic bodies involved in restricting retroviruses such as HIV-1. Here we applied cryogenic correlated light and electron microscopy, combined with electron cryo-tomography, to intact mammalian cells expressing YFP-rhTRIM5α and found the presence of hexagonal nets whose arm lengths were similar to those of the hexagonal nets formed by purified TRIM5α in vitro. We also observed YFP-rhTRIM5α within a diversity of structures with characteristics expected for organelles involved in different stages of macroautophagy, including disorganized protein aggregations (sequestosomes), sequestosomes flanked by flat double-membraned vesicles (sequestosome:phagophore complexes), sequestosomes within double-membraned vesicles (autophagosomes), and sequestosomes within multivesicular autophagic vacuoles (amphisomes or autolysosomes). Vaults were also seen in these structures, consistent with their role in autophagy. Our data 1) support recent reports that TRIM5α can form both well-organized signaling complexes and nonsignaling aggregates, 2) offer images of the macroautophagy pathway in a near-native state, and 3) reveal that vaults arrive early in macroautophagy.
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DOI：
10.1083/jcb.143.7.1883
发表时间：
1998-12-28
期刊：
The Journal of cell biology
影响因子：
--
作者：
Johnston JA;Ward CL;Kopito RR
通讯作者：
Kopito RR
影响因子：
3.3
作者：
Campbell, Edward M.;Dodding, Mark P.;Hope, Thomas J.
通讯作者：
Hope, Thomas J.
影响因子：
3
作者：
Kremer, JR;Mastronarde, DN;McIntosh, JR
通讯作者：
McIntosh, JR
p62/SQSTM1形成自噬降解的蛋白质聚集体，并对亨廷顿蛋白诱导的细胞死亡具有保护作用。
影响因子：
7.8
作者：
Bjorkoy, Geir;Lamark, Trond;Brech, Andreas;Outzen, Heidi;Perander, Maria;Overvatn, Aud;Stenmark, Harald;Johansen, Terje
通讯作者：
Johansen, Terje
在恒河体限制HIV-1时，可视化蛋白酶体的中间体。
影响因子：
7.8
作者：
Campbell, Edward M.;Perez, Omar;Anderson, Jenny L.;Hope, Thomas J.
通讯作者：
Hope, Thomas J.