Redox-dependent Changes in Structure and Function of hHSF1

LIN Zheng*, HUANG Fan, MA Zhong-Fu, XU Kang, LIU Alice Y¹

( The First Hospital Affiliated to Sun Yat-Sen University, Guangzhou, 510080 China; ¹Department of Cell Biology and Neuroscience, Rutgers State University of New Jersey 08854, USA )

Abstract To evaluate the effects of cysteine-SH-directed regents on the redox status, structure and function of human heat shock transcription factor 1 (hHSF1), treatment in vitro of hHSF1 with 0.3-0.5 mmol/L oxidizing reagent diamide (DM) and treatment in vivo of HeLa cells with 1 mmol/L buthionine sulfoximine (BSO), an inhibitor of g-glutamylcysteine synthetase, promoted the formation of a compact, intramolecularly disulfide-crosslinked, stable monomeric form of ox-hHSF1, and blocked the trimerization and activation of HSF1. The effects of diamine were dose-dependent and readily could be completely reversed by adding 0.4-0.5 mmol/L reducing reagent dithiothreitol (DTT) to the samples prior to gel electrophoresis. Computer modeling of the a-helical coiled-coil domains of the HSF1 monomer and trimer showed that the alignment of the N- and C-terminal hydrophobic repeats of HSF1 monomer could bring C3 (Cys153) close to C4 and C5 (Cys373 and Cys378, respectively), in positions permissible for disulfide bond formation under appropriate experimental conditions. The results suggest that redox-dependent thiol-disulfide exchange can provide a mechanism for regulation the conformation and activity of hHSF1.

Key words hHSF1; redox; thiol; cysteine

*Corresponding author: Tel, 86-20-87331142; e-mail, [email protected]