Structure and activation mechanism of the BBSome membrane protein trafficking complex.

Sandeep K Singh,Alan Brown,Fujiet Koh,Matthew Cj Yip,Miao Gui

doi:10.7554/elife.53322

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 β-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.

Highlights

Most eukaryotic cells have a solitary primary cilium capable of sensing both internal and external stimuli (Singla and Reiter, 2006)
Our structures of the BBSome with and without ARL6 show that activation of the BBSome at ciliary membranes requires a swiveling of BBS1bprop that widens a cavity in the body of the BBSome
The breaking of the continuous b-sheet between BBS1 and BBS2 exposes b-edge strands, which are common mediators of protein–protein interactions (Remaut and Waksman, 2006) that have the potential to hydrogen bond to cytosolic regions of transmembrane proteins

Summary

Introduction

Most eukaryotic cells have a solitary primary cilium capable of sensing both internal and external stimuli (Singla and Reiter, 2006). 2013) including those not normally destined for cilia (Datta et al, 2015) This led to a model in which the BBSome promotes retrieval and export of specific transmembrane proteins from the cilium (Nachury, 2018) and the IFT-A complex promotes entry (Mukhopadhyay et al, 2010) (Hirano et al, 2017). We use single-particle cryo-EM to determine structures of the native bovine BBSome complex with and without ARL6 at 3.5 Aand 3.1 Aresolution, respectively These structures allow unambiguous subunit assignment and atomic models to be built for each of the eight BBSome subunits. The structures reveal the mechanism of ARL6-mediated activation and provide new insights into the pathogenesis of BBS-causing mutations and the evolutionary relationship between the BBSome and other transmembrane protein trafficking complexes

Results

Discussion

Materials and methods

Matilda Ziegler Foundation for the Blind