mRNA and protein expression profiling highlights modulation of chromatin-related proteins and chromatin-modifying enzymes in response to HSP90 inhibitors: Identification of PRMT5 as a novel client protein.

LB-326

We used a chemical biology approach involving a combination of mRNA expression profiling and proteomic analysis to screen for genes and proteins that showed altered expression following HSP90 molecular chaperone inhibition. A2780 human ovarian cancer cells were treated with HSP90 inhibitor 17-AAG. cDNA spotted microarrays were used to probe mRNA expression. 2D gel electrophoresis and MALDI mass spectrometry were used to detect altered protein expression. Of particular interest and novelty, were changes in expression of chromatin-associated genes/proteins and chromatin-modifying enzymes. Expression of the heterochromatin protein HP1 was increased, and that of the histone acetyltransferase HAT-1 and the histone arginine methyltransferases PRMT5 after 17-AAG treatment was decreased. The protein expression changes, confirmed by western blotting, were replicated with other active but not inactive HSP90 inhibitors, consistent with an HSP90 mechanism-based on-target effect. Cellular protein acetylation was reduced by 17-AAG, which was shown to have an antagonistic interaction on cell proliferation with the histone deacetylase inhibitor trichostatin A. Immunoprecipitation studies showed that PRMT5 is a HSP90-binding partner and novel potential client protein. This combined mRNA and protein expression profiling chemical biology approach has highlighted a pharmacologically and functionally significant link between chromatin-associated proteins and chromatin-modifying enzymes. It has also identified new genes and proteins that may be involved in response to HSP90 inhibitors and could be potential biomarkers of drug action.