Hsp90 inhibition induces destabilization of actin cytoskeleton in tumor cells: functional significance of Hsp90 interaction with F-actin

Objective: To examine the role of heat shock protein 90 (Hsp90) in the maintenance of actin cytoskeleton in human neuroblastoma tumor cells. Methods: Co-precipitation experiments were performed to examine Hsp90 interaction with actin. Hsp90 and actin interactions were evaluated by protein refolding and acto-myosin motility assays. 17-(Allylamino)-17- demethoxygeldanamycin (17AAG) induced actin cytoskeleton re-organization was examined by laser scanning confocal microcopy. Results: It was shown that inhibition of Hsp90 by 17AAG accelerates detergent induced cell lysis of neuroblastoma tumor cells through destabilization of actin cytoskeleton. The in vitro co-precipitation experiments showed that functional but not mutant Hsp90 binds with F-actin. Among biochemical modifications, phopshorylation and oligomerization enhanced Hsp90 binding with F-actin. F-actin binding to Hsp9O interfered with Hsp90 chaperone activity in protein refolding assays, and Hsp90 binding to F-actin interfered with actin motility on myosin coated flow cell. In the combination treatment, 17AAG irreversibly augmented the effect of cytochalasin D, an inhibitor of actin polymerization. Conclusions: It can be concluded that Hsp90 binds to F-actin in tumor cells and maintains the cellular integrity. The results display a novel element of Hsp90 inhibition in destabilizing the actin cytoskeleton of tumor cells, therefore suggest that 17AAG combination with cytoskeletal disruptor may he effective in combating cancer.