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Structural basis for membrane targeting of the BBSome by ARL6

Nature Structural & Molecular Biology volume 21, pages 10351041 (2014) | Download Citation

Abstract

The BBSome is a coat-like ciliary trafficking complex composed of proteins mutated in Bardet-Biedl syndrome (BBS). A critical step in BBSome-mediated sorting is recruitment of the BBSome to membranes by the GTP-bound Arf-like GTPase ARL6. We have determined crystal structures of Chlamydomonas reinhardtii ARL6–GDP, ARL6–GTP and the ARL6–GTP–BBS1 complex. The structures demonstrate how ARL6–GTP binds the BBS1 β-propeller at blades 1 and 7 and explain why GTP- but not GDP-bound ARL6 can recruit the BBSome to membranes. Single point mutations in the ARL6-GTP-BBS1 interface abolish the interaction of ARL6 with the BBSome and prevent the import of BBSomes into cilia. Furthermore, we show that BBS1 with the M390R mutation, responsible for 30% of all reported BBS disease cases, fails to interact with ARL6–GTP, thus providing a molecular rationale for patient pathologies.

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Acknowledgements

We thank the staff at Swiss Light Source for guidance with X-ray diffraction data collection, the biochemistry core facility and the crystallization facility of the Max Planck Institute of Biochemistry (MPI-B, Munich) for access to crystallization screening and the Bavarian NMR Center for NMR measurement time. We also thank I.B. Schaefer (MPI-B) for DNA encoding BBS subunits, S. Wachter (MPI-B) for assistance with GTPase assays and M. Taschner (MPI-B) for expert advice on protein production from insect cells and for carefully reading the manuscript. This work was funded by an Emmy Noether grant (Deutsche Forschungsgemeinschaft; LO1627/1-1), by the European Research Council (grant 310343) and by the European Molecular Biology Organization Young Investigator program. A.R.N. was supported by the Fayez Sarofim Fellowship of the Damon Runyon Cancer Research Foundation (DRG 2160-13). This work was supported by a grant to M.V.N. from the NIH/National Institute of General Medical Sciences (R01GM089933).

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Affiliations

  1. Department of Structural Cell Biology, Max-Planck-Institute of Biochemistry, Martinsried, Germany.

    • André Mourão
    •  & Esben Lorentzen
  2. Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California, USA.

    • Andrew R Nager
    •  & Maxence V Nachury

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Contributions

A.M. carried out the protein biochemistry and structural biology under the supervision of E.L.; A.R.N. carried out the pulldown experiments of native BBSome with wild-type and mutant ARL6 and the cell biology experiments under the supervision of M.V.N.; A.M. and E.L. designed the experiments and wrote the paper with input from A.R.N. and M.V.N.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Esben Lorentzen.

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https://doi.org/10.1038/nsmb.2920

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