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Golgi apparatus studied in vitreous sections
Author(s) -
BOUCHETMARQUIS C.,
STARKUVIENE V.,
GRABENBAUER M.
Publication year - 2008
Publication title -
journal of microscopy
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.569
H-Index - 111
eISSN - 1365-2818
pISSN - 0022-2720
DOI - 10.1111/j.1365-2818.2008.01988.x
Subject(s) - golgi apparatus , copi , electron microscope , vesicle , organelle , ultrastructure , biophysics , cryo electron microscopy , microbiology and biotechnology , electron tomography , biology , resolution (logic) , secretory pathway , microtubule , endoplasmic reticulum , chemistry , anatomy , membrane , biochemistry , optics , physics , scanning transmission electron microscopy , artificial intelligence , computer science
Summary Cryo‐electron microscopy of vitrified specimen is the method of choice to explore cellular ultrastructure at high resolution as close as possible to the native state and environment. In this study, we investigated the Golgi apparatus – the main organelle of the secretory pathway. Cultured mammalian cells were fixed by high‐pressure freezing, sectioned in vitreous ice and subjected to cryo‐electron microscopy and cryo‐electron tomography. Although the overall morphology of Golgi stacks was comparable to well prepared and plastic‐embedded samples, in detail we reached much higher resolution in terms of distinction between biological structures based on their native density. On cisternal buds and peri‐Golgi vesicles – some associated with microtubules – we detected two different subtypes of COPI coats: (1) a homogenous coat and (2) an inhomogeneous spiky coat, providing an 8–9 nm regularity, clearly distinct from clathrin coat. Next, we monitored the secretion of cargo, namely, procollagen I, through the Golgi complex. Temporally correlated with fluorescence microscopy, we performed three‐dimensional cryo‐electron tomography analysis and detected Golgi cisternae enlarged to saccules, containing cargo and showing inter‐cisternal connections. Our work provides a first step towards the high‐resolution description of the secretory pathway in native vitrified samples and describes the challenges associated with this attempt.