title: HSP90 and its R2TP/Prefoldin-like cochaperone are involved in the cytoplasmic assembly of RNA polymerase II.
abstract: RNA polymerases are key multisubunit cellular enzymes. Microscopy studies indicated that RNA polymerase I assembles near its promoter. However, the mechanism by which RNA polymerase II is assembled from its 12 subunits remains unclear. We show here that RNA polymerase II subunits Rpb1 and Rpb3 accumulate in the cytoplasm when assembly is prevented and that nuclear import of Rpb1 requires the presence of all subunits. Using MS-based quantitative proteomics, we characterized assembly intermediates. These included a cytoplasmic complex containing subunits Rpb1 and Rpb8 associated with the HSP90 cochaperone hSpagh (RPAP3) and the R2TP/Prefoldin-like complex. Remarkably, HSP90 activity stabilized incompletely assembled Rpb1 in the cytoplasm. Our data indicate that RNA polymerase II is built in the cytoplasm and reveal quality-control mechanisms that link HSP90 to the nuclear import of fully assembled enzymes. hSpagh also bound the free RPA194 subunit of RNA polymerase I, suggesting a general role in assembling RNA polymerases.
authors.affiliation: Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Séverine
authors.initials: S
authors.lastname: Boulon
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Bérengère
authors.initials: B
authors.lastname: Pradet-Balade
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Céline
authors.initials: C
authors.lastname: Verheggen
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Dorothée
authors.initials: D
authors.lastname: Molle
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Stéphanie
authors.initials: S
authors.lastname: Boireau
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Marya
authors.initials: M
authors.lastname: Georgieva
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Karim
authors.initials: K
authors.lastname: Azzag
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Marie-Cécile
authors.initials: MC
authors.lastname: Robert
authors.affiliation: Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Yasmeen
authors.initials: Y
authors.lastname: Ahmad
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.
authors.forename: Henry
authors.initials: H
authors.lastname: Neel
authors.affiliation: Wellcome Trust Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, UK.
authors.forename: Angus I
authors.initials: AI
authors.lastname: Lamond
authors.affiliation: Institut de Génétique Moléculaire de Montpellier - UMR 5535 CNRS, Université Montpellier 2, 34293 Montpellier Cedex 5, France.|Université Montpellier 2, 34095 Montpellier Cedex 5, France.|Université Montpellier 1, 34060 Montpellier Cedex 2, France.
authors.forename: Edouard
authors.initials: E
authors.lastname: Bertrand
mesh_terms: D021581:Active Transport, Cell Nucleus / Q000187:drug effects / Q000502:physiology; D053959:Alpha-Amanitin / Q000494:pharmacology; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000378:metabolism*; D045744:Cell Line, Tumor; D003593:Cytoplasm / Q000378:metabolism*; D017930:Genes, Reporter / Q000235:genetics; D015497:HIV-1 / Q000235:genetics; D018841:HSP90 Heat-Shock Proteins / Q000378:metabolism*; D006801:Humans; D018832:Molecular Chaperones / Q000378:metabolism*; D046912:Multiprotein Complexes / Q000187:drug effects / Q000378:metabolism*; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000502:physiology; D025941:Protein Interaction Mapping / Q000379:methods; D055503:Protein Multimerization / Q000187:drug effects / Q000502:physiology*; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D040901:Proteomics; D012318:RNA Polymerase I / Q000378:metabolism; D012319:RNA Polymerase II / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering
chemical_list: D053959:Alpha-Amanitin; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D018841:HSP90 Heat-Shock Proteins; D018832:Molecular Chaperones; D046912:Multiprotein Complexes; D021122:Protein Subunits; D034741:RNA, Small Interfering; C472351:RPAP3 protein, human; C112806:prefoldin; C491783:POLR2C protein, human; D012319:RNA Polymerase II; C410949:RNA polymerase II largest subunit; D012318:RNA Polymerase I