Flexible open conformation of the AP-3 complex explains its role in cargo recruitment at the Golgi.

Autor:	Schoppe J; Department of Biology/Chemistry, Biochemistry Section, Osnabrück University, Osnabrück, Germany., Schubert E; Department of Structural Biochemistry, Max-Planck Institute of Molecular Physiology, Dortmund, Germany., Apelbaum A; Department of Structural Biochemistry, Max-Planck Institute of Molecular Physiology, Dortmund, Germany., Yavavli E; Department of Biology/Chemistry, Biochemistry Section, Osnabrück University, Osnabrück, Germany., Birkholz O; Department of Biology/Chemistry, Biophysics Section, Osnabrück University, Osnabrück, Germany., Stephanowitz H; Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Campus Berlin-Buch, Berlin, Germany., Han Y; Department of Biology/Chemistry, Biochemistry Section, Osnabrück University, Osnabrück, Germany., Perz A; Department of Biology/Chemistry, Biochemistry Section, Osnabrück University, Osnabrück, Germany., Hofnagel O; Department of Structural Biochemistry, Max-Planck Institute of Molecular Physiology, Dortmund, Germany., Liu F; Leibniz-Forschungsinstitut für Molekulare Pharmakologie (FMP), Campus Berlin-Buch, Berlin, Germany., Piehler J; Department of Biology/Chemistry, Biophysics Section, Osnabrück University, Osnabrück, Germany; Center of Cellular Nanoanalytics Osnabrück (CellNanOs), Osnabrück University, Osnabrück, Germany., Raunser S; Department of Structural Biochemistry, Max-Planck Institute of Molecular Physiology, Dortmund, Germany. Electronic address: stefan.raunser@mpi-dortmund.mpg.de., Ungermann C; Department of Biology/Chemistry, Biochemistry Section, Osnabrück University, Osnabrück, Germany; Center of Cellular Nanoanalytics Osnabrück (CellNanOs), Osnabrück University, Osnabrück, Germany. Electronic address: cu@uos.de.
Jazyk:	angličtina
Zdroj:	The Journal of biological chemistry [J Biol Chem] 2021 Nov; Vol. 297 (5), pp. 101334. Date of Electronic Publication: 2021 Oct 22.
DOI:	10.1016/j.jbc.2021.101334
Abstrakt:	Vesicle formation at endomembranes requires the selective concentration of cargo by coat proteins. Conserved adapter protein complexes at the Golgi (AP-3), the endosome (AP-1), or the plasma membrane (AP-2) with their conserved core domain and flexible ear domains mediate this function. These complexes also rely on the small GTPase Arf1 and/or specific phosphoinositides for membrane binding. The structural details that influence these processes, however, are still poorly understood. Here we present cryo-EM structures of the full-length stable 300 kDa yeast AP-3 complex. The structures reveal that AP-3 adopts an open conformation in solution, comparable to the membrane-bound conformations of AP-1 or AP-2. This open conformation appears to be far more flexible than AP-1 or AP-2, resulting in compact, intermediate, and stretched subconformations. Mass spectrometrical analysis of the cross-linked AP-3 complex further indicates that the ear domains are flexibly attached to the surface of the complex. Using biochemical reconstitution assays, we also show that efficient AP-3 recruitment to the membrane depends primarily on cargo binding. Once bound to cargo, AP-3 clustered and immobilized cargo molecules, as revealed by single-molecule imaging on polymer-supported membranes. We conclude that its flexible open state may enable AP-3 to bind and collect cargo at the Golgi and could thus allow coordinated vesicle formation at the trans-Golgi upon Arf1 activation. Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article. (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)
Databáze:	MEDLINE
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