Structure and activation mechanism of the BBSome membrane protein trafficking complex
- Harvard Medical School, United States
Abstract
Bardet-Biedl syndrome (BBS) is a currently incurable ciliopathy caused by the failure to correctly establish or maintain cilia-dependent signaling pathways. Eight proteins associated with BBS assemble into the BBSome, a key regulator of the ciliary membrane proteome. We report the electron cryomicroscopy (cryo-EM) structures of the native bovine BBSome in inactive and active states at 3.1 and 3.5 Å resolution, respectively. In the active state, the BBSome is bound to an Arf-family GTPase (ARL6/BBS3) that recruits the BBSome to ciliary membranes. ARL6 recognizes a composite binding site formed by BBS1 and BBS7 that is occluded in the inactive state. Activation requires an unexpected swiveling of the b-propeller domain of BBS1, the subunit most frequently implicated in substrate recognition, which widens a central cavity of the BBSome. Structural mapping of disease-causing mutations suggests that pathogenesis results from folding defects and the disruption of autoinhibition and activation.
Data availability
The EM density map for the BBSome has been deposited under accession code EMD-21144 and the EM density map for the BBSome:ARL6:GTP complex has been deposited under accession code EMD-21145. Masks and maps from multibody refinement are included as additional maps in these depositions. The corresponding atomic models have been deposited under accession codes 6VBU and 6VBV.
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Structure of the bovine BBSome (map)Electron Microscopy Data Bank, EMD-21144.
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Structure of the bovine BBSome:ARL6:GTP complex (map)Electron Microscopy Data Bank, EMD-21145.
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Structure of the bovine BBSome:ARL6:GTP complex (model)Protein Data Bank, 6VBV.
Article and author information
Author details
Funding
Pew Charitable Trusts
- Alan Brown
Internal Retinal Research Foundation
- Alan Brown
E. Matilda Ziegler Foundation for the Blind
- Alan Brown
Richard and Susan Smith Family Foundation
- Alan Brown
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Andrew P Carter, MRC Laboratory of Molecular Biology, United Kingdom
Version history
- Received: November 4, 2019
- Accepted: January 6, 2020
- Accepted Manuscript published: January 15, 2020 (version 1)
- Version of Record published: February 13, 2020 (version 2)
Copyright
© 2020, Singh 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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Structure and activation mechanism of the BBSome membrane protein trafficking complex
eLife 9:e53322.
https://doi.org/10.7554/eLife.53322
Further reading
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- Structural Biology and Molecular Biophysics
The structures of the bovine and human BBSome reveal that a conformational change is required to recruit the complex to the ciliary membrane.
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- Structural Biology and Molecular Biophysics
The proteasome controls levels of most cellular proteins, and its activity is regulated under stress, quiescence, and inflammation. However, factors determining the proteasomal degradation rate remain poorly understood. Proteasome substrates are conjugated with small proteins (tags) like ubiquitin and Fat10 to target them to the proteasome. It is unclear if the structural plasticity of proteasome-targeting tags can influence substrate degradation. Fat10 is upregulated during inflammation, and its substrates undergo rapid proteasomal degradation. We report that the degradation rate of Fat10 substrates critically depends on the structural plasticity of Fat10. While the ubiquitin tag is recycled at the proteasome, Fat10 is degraded with the substrate. Our results suggest significantly lower thermodynamic stability and faster mechanical unfolding in Fat10 compared to ubiquitin. Long-range salt bridges are absent in the Fat10 structure, creating a plastic protein with partially unstructured regions suitable for proteasome engagement. Fat10 plasticity destabilizes substrates significantly and creates partially unstructured regions in the substrate to enhance degradation. NMR-relaxation-derived order parameters and temperature dependence of chemical shifts identify the Fat10-induced partially unstructured regions in the substrate, which correlated excellently to Fat10-substrate contacts, suggesting that the tag-substrate collision destabilizes the substrate. These results highlight a strong dependence of proteasomal degradation on the structural plasticity and thermodynamic properties of the proteasome-targeting tags.
