Single-molecule analysis reveals self assembly and nanoscale segregation of two distinct cavin subcomplexes on caveolae
Yann Gambin,
Nicholas Ariotti,
Kerrie-Ann McMahon,
Michele Bastiani,
Emma Sierecki,
Oleksiy Kovtun,
Mark E Polinkovsky,
Astrid Magenau,
WooRam Jung,
Satomi Okano,
Yong Zhou,
Natalya Leneva,
Sergey Mureev,
Wayne Johnston,
Katharina Gaus,
John F Hancock,
Brett M Collins,
Kirill Alexandrov,
Robert G Parton
Affiliations
Yann Gambin
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Nicholas Ariotti
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Kerrie-Ann McMahon
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Michele Bastiani
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Emma Sierecki
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Oleksiy Kovtun
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Mark E Polinkovsky
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Astrid Magenau
Center for Vascular Research, University of New South Wales, Sydney, Australia; Australian Centre for Nanomedicine, University of New South Wales, Sydney, Australia
WooRam Jung
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Satomi Okano
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Yong Zhou
Department of Integrative Biology and Pharmacology, The University of Texas Medical School at Houston, Houston, United States
Natalya Leneva
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Sergey Mureev
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Wayne Johnston
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Katharina Gaus
Center for Vascular Research, University of New South Wales, Sydney, Australia; Australian Centre for Nanomedicine, University of New South Wales, Sydney, Australia
John F Hancock
Department of Integrative Biology and Pharmacology, The University of Texas Medical School at Houston, Houston, United States
Brett M Collins
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Kirill Alexandrov
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia
Robert G Parton
Institute for Molecular Bioscience, The University of Queensland, Brisbane, Australia; Centre for Microscopy and Microanalysis, The University of Queensland, Brisbane, Australia
In mammalian cells three closely related cavin proteins cooperate with the scaffolding protein caveolin to form membrane invaginations known as caveolae. Here we have developed a novel single-molecule fluorescence approach to directly observe interactions and stoichiometries in protein complexes from cell extracts and from in vitro synthesized components. We show that up to 50 cavins associate on a caveola. However, rather than forming a single coat complex containing the three cavin family members, single-molecule analysis reveals an exquisite specificity of interactions between cavin1, cavin2 and cavin3. Changes in membrane tension can flatten the caveolae, causing the release of the cavin coat and its disassembly into separate cavin1-cavin2 and cavin1-cavin3 subcomplexes. Each of these subcomplexes contain 9 ± 2 cavin molecules and appear to be the building blocks of the caveolar coat. High resolution immunoelectron microscopy suggests a remarkable nanoscale organization of these separate subcomplexes, forming individual striations on the surface of caveolae.
