title: Subunit connectivity, assembly determinants and architecture of the yeast exocyst complex.
abstract: The exocyst is a hetero-octameric complex that has been proposed to serve as the tethering complex for exocytosis, although it remains poorly understood at the molecular level. Here, we purified endogenous exocyst complexes from Saccharomyces cerevisiae and showed that they are stable and consist of all eight subunits with equal stoichiometry. Using a combination of biochemical and auxin induced-degradation experiments in yeast, we mapped the subunit connectivity, identified two stable four-subunit modules within the octamer and demonstrated that several known exocyst-binding partners are not necessary for exocyst assembly and stability. Furthermore, we visualized the structure of the yeast complex by using negative-stain electron microscopy; our results indicate that the exocyst exists predominantly as a stable, octameric complex with an elongated architecture that suggests that the subunits are contiguous helical bundles packed together into a bundle of long rods.
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Margaret R
authors.initials: MR
authors.lastname: Heider
authors.affiliation: Department of Biochemistry, University of Utah, Salt Lake City, Utah, USA.
authors.forename: Mingyu
authors.initials: M
authors.lastname: Gu
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Caroline M
authors.initials: CM
authors.lastname: Duffy
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Anne M
authors.initials: AM
authors.lastname: Mirza
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Laura L
authors.initials: LL
authors.lastname: Marcotte
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Alexandra C
authors.initials: AC
authors.lastname: Walls
authors.affiliation: Department of Biochemistry, University of Utah, Salt Lake City, Utah, USA.
authors.forename: Nicholas
authors.initials: N
authors.lastname: Farrall
authors.affiliation: Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, New York, USA.
authors.forename: Zhanna
authors.initials: Z
authors.lastname: Hakhverdyan
authors.affiliation: Division of Biological Chemistry and Drug Discovery, University of Dundee, Dundee, UK.
authors.forename: Mark C
authors.initials: MC
authors.lastname: Field
authors.affiliation: Laboratory of Cellular and Structural Biology, The Rockefeller University, New York, New York, USA.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Rout
authors.affiliation: Department of Biochemistry, University of Utah, Salt Lake City, Utah, USA.|Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, California, USA.
authors.forename: Adam
authors.initials: A
authors.lastname: Frost
authors.affiliation: Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts, USA.
authors.forename: Mary
authors.identifier: http://orcid.org/0000-0003-2297-8053
authors.initials: M
authors.lastname: Munson
mesh_terms: D005089:Exocytosis; D046911:Macromolecular Substances / Q000737:chemistry* / Q000302:isolation & purification* / Q000648:ultrastructure; D046529:Microscopy, Electron, Transmission; D011485:Protein Binding; D055550:Protein Stability; D020836:Protein Structure, Quaternary; D012441:Saccharomyces cerevisiae / Q000502:physiology*; D033921:Vesicular Transport Proteins / Q000737:chemistry* / Q000302:isolation & purification* / Q000648:ultrastructure
chemical_list: D046911:Macromolecular Substances; D033921:Vesicular Transport Proteins