RT Journal Article
SR Electronic
T1 Chaperone Activity and Dimerization Properties of Hsp90<em>α</em> and Hsp90<em>β</em> in Glucocorticoid Receptor Activation by the Multiprotein Hsp90/Hsp70-Dependent Chaperone Machinery
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 984
OP 991
DO 10.1124/mol.118.112516
VO 94
IS 3
A1 Yoshihiro Morishima
A1 Ranjit K. Mehta
A1 Miyako Yoshimura
A1 Miranda Lau
A1 Daniel R. Southworth
A1 Theodore S. Lawrence
A1 William B. Pratt
A1 Mukesh K. Nyati
A1 Yoichi Osawa
YR 2018
UL http://molpharm.aspetjournals.org/content/94/3/984.abstract
AB Several hundred proteins cycle into heterocomplexes with a dimer of the chaperone heat shock protein 90 (Hsp90), regulating their activity and turnover. There are two isoforms of Hsp90, Hsp90α and Hsp90β, and their relative chaperone activities and composition in these client protein•Hsp90 heterocomplexes has not been determined. Here, we examined the activity of human Hsp90α and Hsp90β in a purified five-protein chaperone machinery that assembles glucocorticoid receptor (GR)•Hsp90 heterocomplexes to generate high-affinity steroid-binding activity. We found that human Hsp90α and Hsp90β have equivalent chaperone activities, and when mixed together in this assay, they formed only GR•Hsp90αα and GR•Hsp90ββ homodimers and no GR•Hsp90αβ heterodimers. In contrast, GR•Hsp90 heterocomplexes formed in human embryonic kidney (HEK) cells also contain GR•Hsp90αβ heterodimers. The formation of GR•Hsp90αβ heterodimers in HEK cells probably reflects the longer time permitted for exchange to form Hsp90αβ heterodimers in the cell versus in the cell-free assembly conditions. This purified GR-activating chaperone machinery can be used to determine how modifications of Hsp90 affect its chaperone activity. To that effect, we have tested whether the unique phosphorylation of Hsp90α at threonines 5 and 7 that occurs during DNA damage repair affects its chaperone activity. We showed that the phosphomimetic mutant Hsp90α T5/7D has the same intrinsic chaperone activity as wild-type human Hsp90α in activation of GR steroid-binding activity by the five-protein machinery, supporting the conclusion that T5/7 phosphorylation does not affect Hsp90α chaperone activity.