A mammalian system for high-resolution imaging of intact cells by cryo-electron tomography

被引：2
作者：
Li, Xia [1 ,2 ,3 ,4 ]
Park, Donghyun [1 ,2 ]
Chang, Yunjie [1 ,2 ]
Radhakrishnan, Abhijith [5 ]
Wu, Hangjun [6 ,7 ]
Wang, Pei [8 ]
Liu, Jun [1 ,2 ]
机构：
[1] Yale Sch Med, Dept Microbial Pathogenesis, New Haven, CT 06516 USA
[2] Yale Sch Med, Microbial Sci Inst, New Haven, CT 06516 USA
[3] Nantong Univ, Inst Special Environm Med, Nantong 226000, Jiangsu, Peoples R China
[4] Nantong Univ, Coinnovat Ctr Neuroregenerat, Nantong 226000, Jiangsu, Peoples R China
[5] Yale Sch Med, Dept Cell Biol, New Haven, CT 06520 USA
[6] Zhejiang Univ, Sir Run Run Shaw Hosp, Sch Med, Ctr Cryo Electron Microscopy, Hangzhou 310058, Zhejiang, Peoples R China
[7] Zhejiang Univ, Sir Run Run Shaw Hosp, Sch Med, Dept Pathol, Hangzhou 310058, Zhejiang, Peoples R China
[8] Chinese Acad Sci, Inst Biophys, Beijing 100101, Peoples R China
来源：
PROGRESS IN BIOPHYSICS & MOLECULAR BIOLOGY | 2021年 / 160卷
基金：
美国国家卫生研究院; 中国国家自然科学基金;
关键词：
Mammalian cells; In situ structure; Molecular machine; Organelle; PC12(+) cells; NERVE GROWTH-FACTOR; MOLECULAR-ORGANIZATION; ELECTRON-MICROSCOPY; CRYO-FLUORESCENCE; EUKARYOTIC CELLS; ER MEMBRANES; PC12; CELLS; DIFFERENTIATION; ARCHITECTURE; SYNAPSES;
D O I：
10.1016/j.pbiomolbio.2020.09.005
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
Mammalian cells contain an elaborate network of organelles and molecular machines that orchestrate essential cellular processes. Visualization of this network at a molecular level is vital for understanding these cellular processes. Here we present a model system based on nerve growth factor (NGF)-differentiated PC12 cells (PC12(+)) and suitable for high resolution imaging of organelles and molecular machines in situ. We detail an optimized imaging pipeline that effectively combines correlative light and electron microscopy (CLEM), cryo-focused ion beam (cryo-FIB), cryo-electron tomography (cryo-ET), and sub-tomogram averaging to produce three-dimensional and molecular resolution snapshots of organelles and molecular machines in near-native cellular environments. Our studies demonstrate that cryoET imaging of PC12(+) systems provides an accessible and highly efficient avenue for dissecting specific cellular processes in mammalian cells at high resolution. (C) 2020 Elsevier Ltd. All rights reserved.
引用
收藏
页码：87 / 96
页数：10
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