Geldanamycin Restores a Defective Heat Shock Responsein Vivo

Abstract

Induced expression of heat shock proteins (Hsps) plays a central role in promoting cellular survival after environmental and physiological stress. We have previously shown that scrapie-infected mouse neuroblastoma (ScN2a) cells fail to induce the expression of Hsp72 and Hsp28 after various stress conditions. Here we present evidence that this impaired stress response is due to an altered regulation of HSF1 activity. Upon stress in ScN2a cells, HSF1 was converted into hyperphosphorylated trimers but failed to acquire transactivation competence. A kinetic analysis of HSF1 activation revealed that in ScN2a cells trimer formation after stress was efficient, but disassembly of trimers proceeded much faster than in the uninfected cell line. Geldanamycin, a Hsp90-binding drug, significantly delayed disassembly of HSF1 trimers after a heat shock and restored stressinduced expression of Hsp72 in ScN2a cells. Heat-induced Hsp72 expression required geldanamycin to be present; following removal of the drug ScN2a cells again lost their ability to mount a stress response. Thus, our studies show that a defective stress response can be pharmacologically restored and suggest that the HSF1 deactivation pathway may play an important role in the regulation of Hsp expression.

Up-regulation of Heat Shock Protein 27 Induces Resistance to 17-Allylamino-Demethoxygeldanamycin through a Glutathione-Mediated Mechanism

“…Previous studies have shown that Hsp27 is highly up-regulated after treatment of cells with Hsp90-directed agents such as 17-AAG (10,12). This prompted us to hypothesize that up-regulation of Hsp27 would decrease 17-AAG cell sensitivity and contribute to 17-AAG resistance.…”

“…Because these proteins are dramatically up-regulated in response to treatment with Hsp90 inhibitors such as GA or 17-AAG (10)(11)(12), their potential involvement in resistance to Hsp90 inhibitors in the clinic has recently gained interest (16). Other studies have shown the protection of cancer cells from 17-AAG-induced death by Hsp70, which is known to be up-regulated in response to Hsp90-directed agents (17).…”

“…Hsp27 has also been identified as a participant in cellular chemoresistance in response to treatment with anticancer drugs such as cisplatin (19) and has been shown to be up-regulated in response to GA and 17-AAG (10)(11)(12). The conserved a-crystallin domain that characterizes the small heat shock protein family is found in the COOH terminus of Hsp27 and serves to mediate protein-protein interactions that govern the formation of high molecular weight oligomers (14,20).…”

“…GA derivatives such as 17-AAG and 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG), which have similar activity to GA, are now in phase I and phase II clinical trials (5)(6)(7)(8). Treatment with these compounds produces a variety of responses in cells, including inhibition of Hsp90 chaperone complex activity (1), degradation of client proteins (9), and up-regulation of proteins such as Hsp70 and Hsp27 (10)(11)(12).…”

Post-translational Import of the Prion Protein into the Endoplasmic Reticulum Interferes with Cell Viability

“…Yeast cells were transformed using the LiCl method (36,37). N2a cells were transfected using the LipofectAMINE Plus reagent (Invitrogen) as described previously (38).…”