Targeting of the protein chaperone, HSP90, by the transformation suppressing agent, radicicol

Abstract

Radicicol, a macrocyclic anti-fungal antibiotic, has the ability to suppress transformation by diverse oncogenes such as Src, Ras and Mos. Despite this useful property, the mechanism by which radicicol exerts its antitransformation e ects is currently unknown. To understand the transformation-suppressing e ects of radicicol, a biotinylated derivative of radicicol was chemically synthesized and used as a probe in a Western-blot format to visualize cellular proteins that interact with radicicol. In transformed and untransformed mouse ®broblasts, the most prominent cellular protein that bound to radicicol had a molecular weight of approximately 90 kDa. Further analysis revealed that this protein was the mouse homologue of the 90 kDa heat shock protein (HSP90). This was con®rmed by demonstrating the ability of radicicol to speci®cally bind puri®ed human HSP90. Speci®city of binding was demonstrated by the inhibition of binding of biotinylated radicicol by the native drug. Taken together with other studies the present observations suggest that the anti-transformation e ects of radicicol may be mediated, at least in part, by the association of radicicol with HSP90 and the consequent dissociation of the Raf/HSP90 complex leading to the attenuation of the Ras/MAP kinase signal transduction pathway.

Selective apoptosis of tandemly duplicated FLT3-transformed leukemia cells by Hsp90 inhibitors

“…Radicicol, which was developed as an anti-fungal agent and reversed the tumor phenotype of Srcand Ras-transformed cell lines, also targets Hsp90. 31,32 We thus speculated that Hsp90 was associated with the selective sensitivity of TDFLT3 to HA, although the interaction between FLT3 and Hsp90 was unknown.…”

Regulation of PIDD auto-proteolysis and activity by the molecular chaperone Hsp90

“…In a fashion similar to other Hsp90 client proteins like IRF-1 (interferon regulators factor-1), 16 both GA and Radicicol (an alternative Hsp90 inhibitor 17 ) treatments led to a dramatic decrease of PIDD protein levels ( Figure 2a and Supplementary Figure 2A, respectively). Noteworthy, the half-life of PIDD was around 2 h during GA treatment, whereas other Hsp90 client such as IKKa and RIP1 18,19 were not markedly affected after 8 h of treatment (Figure 2a and Supplementary Figure 2B).…”

Hsp90 Is Essential for the Synthesis and Subsequent Membrane Association, But Not the Maintenance, of the Src-Kinase p56lck

“…SupT1 cells were metabolically labeled with [ 35 S]methionine/cysteine for 3.5 h in the presence or absence of the Hsp90 inhibitors geldanamycin and radicicol. Both of these agents readily cross cellular membranes and bind with high affinity to Hsp90, thereby blocking the activity of this protein (Whitesell et al, 1994;Schulte et al, 1998;Sharma et al, 1998). After labeling, cells were lysed and analyzed by immunoprecipitation for newly synthesized Lck and by Western blotting for total Lck.…”

“…These studies have been facilitated by the use of two specific inhibitors of Hsp90 activity, geldanamycin (Whitesell et al, 1994) and radicicol (Schulte et al, 1998;Sharma et al, 1998). These naturally occurring drugs were initially identified as antitumor agents and inhibitors of tyrosine kinase activity.…”