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Quantitative proteomics of the yeast Hsp70/Hsp90 interactomes during DNA damage reveal chaperone-dependent regulation of ribonucleotide reductase.

Abstract：

UNLABELLED:The highly conserved molecular chaperones Hsp90 and Hsp70 are indispensible for folding and maturation of a significant fraction of the proteome, including many proteins involved in signal transduction and stress response. To examine the dynamics of chaperone-client interactions after DNA damage, we applied quantitative affinity-purification mass spectrometry (AP-MS) proteomics to characterize interactomes of the yeast Hsp70 isoform Ssa1 and Hsp90 isoform Hsp82 before and after exposure to methyl methanesulfonate. Of 256 proteins identified and quantified via (16)O(/18)O labeling and LC-MS/MS, 142 are novel Hsp70/90 interactors. Nearly all interactions remained unchanged or decreased after DNA damage, but 5 proteins increased interactions with Ssa1 and/or Hsp82, including the ribonucleotide reductase (RNR) subunit Rnr4. Inhibiting Hsp70 or 90 chaperone activity destabilized Rnr4 in yeast and its vertebrate homolog hRMM2 in breast cancer cells. In turn, pre-treatment of cancer cells with chaperone inhibitors sensitized cells to the RNR inhibitor gemcitabine, suggesting a novel chemotherapy strategy. All MS data have been deposited in the ProteomeXchange with identifier PXD001284. BIOLOGICAL SIGNIFICANCE:This study provides the dynamic interactome of the yeast Hsp70 and Hsp90 under DNA damage which suggest key roles for the chaperones in a variety of signaling cascades. Importantly, the cancer drug target ribonucleotide reductase was shown to be a client of Hsp70 and Hsp90 in both yeast and breast cancer cells. As such, this study highlights the potential of a novel cancer therapeutic strategy that exploits the synergy of chaperone and ribonucleotide reductase inhibitors.

journal_name

J Proteomics

journal_title

Journal of proteomics

authors

Truman AW,Kristjansdottir K,Wolfgeher D,Ricco N,Mayampurath A,Volchenboum SL,Clotet J,Kron SJ

doi

10.1016/j.jprot.2014.09.028

subject

Has Abstract

pub_date

2015-01-01 00:00:00

pages

285-300

eissn

1874-3919

issn

1876-7737

pii

S1874-3919(14)00461-8

journal_volume

112

pub_type

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Quantitative proteomics of the yeast Hsp70/Hsp90 interactomes during DNA damage reveal chaperone-dependent regulation of ribonucleotide reductase.