HSP90 and its R2TP/Prefoldin-like cochaperone are involved in the cytoplasmic assembly of RNA polymerase II

Séverine Boulon; Bérengère Pradet-Balade; Céline Verheggen; Dorothée Molle; Stéphanie Boireau; Marya Georgieva; Karim Azzag; Marie Cécile Robert; Yasmeen Ahmad; Henry Neel; Angus I. Lamond; Edouard Bertrand

doi:10.1016/j.molcel.2010.08.023

HSP90 and its R2TP/Prefoldin-like cochaperone are involved in the cytoplasmic assembly of RNA polymerase II

Séverine Boulon, Bérengère Pradet-Balade, Céline Verheggen, Dorothée Molle, Stéphanie Boireau, Marya Georgieva, Karim Azzag, Marie Cécile Robert, Yasmeen Ahmad, Henry Neel, Angus I. Lamond, Edouard Bertrand

Research output: Contribution to journal › Article › peer-review

218 Scopus citations

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.

Original language	English (US)
Pages (from-to)	912-924
Number of pages	13
Journal	Molecular Cell
Volume	39
Issue number	6
DOIs	https://doi.org/10.1016/j.molcel.2010.08.023
State	Published - Sep 2010
Externally published	Yes

Bibliographical note

Funding Information:
We thank M. Kress for the gift of DsRed2-p54/RCK and the idea of using it to target fusion proteins to P-bodies. We thank M. Vidal for the gift of the human ORFeome 3.1 and U. Rothbauer and H. Leonhardt for the gift of GFP-TRAP beads. We thank R. Bordonné for critical reading of the manuscript. A.I.L. is a Wellcome Trust Principal Research Fellow. S. Boulon is a long-term fellow of the Human Frontier Science Program (HFSP), and D.M. had a fellowship from ANRS. This work was supported by grants from SIDACTION; ANRS; La Ligue Contre le Cancer; ARC; ANR; Eurasnet; PROSPECTS (PROteomics SPECification in Time and Space), which stems from the European Commission's FP7 (GA HEALTH-F4-2008-201648); and RASOR (Radical Solutions for Researching the proteome).

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@article{362d4dbe220842228864dbd8c2e80d16,

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.",

author = "S{\'e}verine Boulon and B{\'e}reng{\`e}re Pradet-Balade and C{\'e}line Verheggen and Doroth{\'e}e Molle and St{\'e}phanie Boireau and Marya Georgieva and Karim Azzag and Robert, {Marie C{\'e}cile} and Yasmeen Ahmad and Henry Neel and Lamond, {Angus I.} and Edouard Bertrand",

note = "Funding Information: We thank M. Kress for the gift of DsRed2-p54/RCK and the idea of using it to target fusion proteins to P-bodies. We thank M. Vidal for the gift of the human ORFeome 3.1 and U. Rothbauer and H. Leonhardt for the gift of GFP-TRAP beads. We thank R. Bordonn{\'e} for critical reading of the manuscript. A.I.L. is a Wellcome Trust Principal Research Fellow. S. Boulon is a long-term fellow of the Human Frontier Science Program (HFSP), and D.M. had a fellowship from ANRS. This work was supported by grants from SIDACTION; ANRS; La Ligue Contre le Cancer; ARC; ANR; Eurasnet; PROSPECTS (PROteomics SPECification in Time and Space), which stems from the European Commission's FP7 (GA HEALTH-F4-2008-201648); and RASOR (Radical Solutions for Researching the proteome). ",

year = "2010",

month = sep,

doi = "10.1016/j.molcel.2010.08.023",

language = "English (US)",

volume = "39",

pages = "912--924",

journal = "Molecular Cell",

issn = "1097-2765",

publisher = "Cell Press",

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T1 - HSP90 and its R2TP/Prefoldin-like cochaperone are involved in the cytoplasmic assembly of RNA polymerase II

AU - Boulon, Séverine

AU - Pradet-Balade, Bérengère

AU - Verheggen, Céline

AU - Molle, Dorothée

AU - Boireau, Stéphanie

AU - Georgieva, Marya

AU - Azzag, Karim

AU - Robert, Marie Cécile

AU - Ahmad, Yasmeen

AU - Neel, Henry

AU - Lamond, Angus I.

AU - Bertrand, Edouard

N1 - Funding Information: We thank M. Kress for the gift of DsRed2-p54/RCK and the idea of using it to target fusion proteins to P-bodies. We thank M. Vidal for the gift of the human ORFeome 3.1 and U. Rothbauer and H. Leonhardt for the gift of GFP-TRAP beads. We thank R. Bordonné for critical reading of the manuscript. A.I.L. is a Wellcome Trust Principal Research Fellow. S. Boulon is a long-term fellow of the Human Frontier Science Program (HFSP), and D.M. had a fellowship from ANRS. This work was supported by grants from SIDACTION; ANRS; La Ligue Contre le Cancer; ARC; ANR; Eurasnet; PROSPECTS (PROteomics SPECification in Time and Space), which stems from the European Commission's FP7 (GA HEALTH-F4-2008-201648); and RASOR (Radical Solutions for Researching the proteome).

PY - 2010/9

Y1 - 2010/9

N2 - 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.

AB - 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.

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U2 - 10.1016/j.molcel.2010.08.023

DO - 10.1016/j.molcel.2010.08.023

M3 - Article

C2 - 20864038

AN - SCOPUS:77956898581

SN - 1097-2765

VL - 39

SP - 912

EP - 924

JO - Molecular Cell

JF - Molecular Cell

IS - 6

ER -

HSP90 and its R2TP/Prefoldin-like cochaperone are involved in the cytoplasmic assembly of RNA polymerase II

Abstract

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