The structure of the COPI coat determined within the cell
Abstract
COPI-coated vesicles mediate trafficking within the Golgi apparatus and from the Golgi to the endoplasmic reticulum. The structures of membrane protein coats, including COPI, have been extensively studied with in vitro reconstitution systems using purified components. In a previous paper (Dodonova et al., 2017), we determined a complete structural model of the in vitro reconstituted COPI coat. Here, we applied cryo-focused ion beam milling, cryo-electron tomography and subtomogram averaging to determine the native structure of the COPI coat within vitrified Chlamydomonas reinhardtii cells. The native algal structure resembles the in vitro mammalian structure, but additionally reveals cargo bound beneath β'-COP. We find that all coat components disassemble simultaneously and relatively rapidly after budding. Structural analysis in situ, maintaining Golgi topology, shows that vesicles change their size, membrane thickness, and cargo content as they progress from cis to trans, but the structure of the coat machinery remains constant.
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Funding
Deutsche Forschungsgemeinschaft (SFB1129 (Z2))
- John AG Briggs
Deutsche Forschungsgemeinschaft (SFB-1035/Project A01)
- Wolfgang Baumeister
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Margaret S. Robinson, University of Cambridge, United Kingdom
Version history
- Received: October 4, 2017
- Accepted: November 16, 2017
- Accepted Manuscript published: November 17, 2017 (version 1)
- Version of Record published: December 5, 2017 (version 2)
Copyright
© 2017, Bykov et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Further reading
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- Structural Biology and Molecular Biophysics
- Cell Biology
Advances in imaging techniques have shed new light on the structure of vesicles formed by COPI protein complexes.
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- Structural Biology and Molecular Biophysics
The physical basis of phase separation is thought to consist of the same types of bonds that specify conventional macromolecular interactions yet is unsatisfyingly often referred to as ‘fuzzy’. Gaining clarity on the biogenesis of membraneless cellular compartments is one of the most demanding challenges in biology. Here, we focus on the chromosome passenger complex (CPC), that forms a chromatin body that regulates chromosome segregation in mitosis. Within the three regulatory subunits of the CPC implicated in phase separation – a heterotrimer of INCENP, Survivin, and Borealin – we identify the contact regions formed upon droplet formation using hydrogen/deuterium exchange mass spectrometry (HXMS). These contact regions correspond to some of the interfaces seen between individual heterotrimers within the crystal lattice they form. A major contribution comes from specific electrostatic interactions that can be broken and reversed through initial and compensatory mutagenesis, respectively. Our findings reveal structural insight for interactions driving liquid-liquid demixing of the CPC. Moreover, we establish HXMS as an approach to define the structural basis for phase separation.