Letter | Published:

Functional architecture of the retromer cargo-recognition complex

Nature volume 449, pages 10631067 (25 October 2007) | Download Citation

Abstract

The retromer complex1,2 is required for the sorting of acid hydrolases to lysosomes3,4,5,6,7, transcytosis of the polymeric immunoglobulin receptor8, Wnt gradient formation9,10, iron transporter recycling11 and processing of the amyloid precursor protein12. Human retromer consists of two smaller complexes: the cargo recognition VPS26–VPS29–VPS35 heterotrimer and a membrane-targeting heterodimer or homodimer of SNX1 and/or SNX2 (ref. 13). Here we report the crystal structure of a VPS29–VPS35 subcomplex showing how the metallophosphoesterase-fold subunit VPS29 (refs 14, 15) acts as a scaffold for the carboxy-terminal half of VPS35. VPS35 forms a horseshoe-shaped, right-handed, α-helical solenoid, the concave face of which completely covers the metal-binding site of VPS29, whereas the convex face exposes a series of hydrophobic interhelical grooves. Electron microscopy shows that the intact VPS26–VPS29–VPS35 complex is a stick-shaped, flexible structure, approximately 21 nm long. A hybrid structural model derived from crystal structures, electron microscopy, interaction studies and bioinformatics shows that the α-solenoid fold extends the full length of VPS35, and that VPS26 is bound at the opposite end from VPS29. This extended structure presents multiple binding sites for the SNX complex and receptor cargo, and appears capable of flexing to conform to curved vesicular membranes.

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Acknowledgements

We thank H. Shi for contributing to the early stages of this project; X. Zhu and H. T. Tsai for technical assistance; B. Canagarajah for bioinformatics and computational crystallography support; E. Tyler for artwork; Z.-Q. Fu and all the staff of APS SER-CAT beamline 22 for assistance with data collection; and C. Haft for antibodies to retromer. Use of the APS was supported by the US DOE, Basic Energy Sciences, Office of Science. This project was funded by the Intramural Programs of NIDDK, NICHD and NIAMS, NIH.

Author Contributions A.L.R. and A.H. expressed and purified protein complexes, crystallized the VPS29–VPS35 C-terminal subcomplex, collected crystallographic data, and determined and refined the crystal structure; A.H. carried out phosphatase assays; R.R. and N.M. carried out immunoprecipitation and optical microscopy studies; G.E. and A.C.S. carried out and interpreted electron microscopy studies; A.V.K. carried out sequence analysis; and J.H.H., J.S.B. and A.C.S. designed the study. A.H. and A.L.R. contributed equally to this study.

Crystallographic coordinates have been deposited with the Protein Data Bank with accession number 2R17.

Author information

Author notes

    • Aitor Hierro
    •  & Adriana L. Rojas

    These authors contributed equally to this work.

Affiliations

  1. Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases,

    • Aitor Hierro
    • , Adriana L. Rojas
    •  & James H. Hurley
  2. Cell Biology and Metabolism Branch, National Institute of Child Health and Human Development, and,

    • Raul Rojas
    • , Namita Murthy
    •  & Juan S. Bonifacino
  3. Laboratory of Structural Biology, National Institute of Arthritis, Musculoskeletal, and Skin Diseases, National Institutes of Health, US Department of Health and Human Services, Bethesda, Maryland 20892, USA

    • Grégory Effantin
    •  & Alasdair C. Steven
  4. Centre de Recherches de Biochimie Macromoléculaire, CNRS, University of Montpellier, 1919 Route de Mende, 34293 Montpellier, France

    • Andrey V. Kajava

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Competing interests

Crystallographic coordinates have been deposited with the Protein Data Bank with accession number 2R17. Reprints and permissions information is available at www.nature.com/reprints.

Corresponding author

Correspondence to James H. Hurley.

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https://doi.org/10.1038/nature06216

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