Analysis of different methylation patterns of histone H3 between androgen-dependent and androgen-independent prostate cancer cells by heavy methyl SILAC

AIM：To study the epigenetic mechanisms involved in the evolution of prostate cancer from an androgen-dependent state to an androgen-independent state, and the global difference of histone H3 methylation between androgen-dependent and -independent prostate cancer cells. METHODS：The methylation sites and patterns of histone H3 in androgen-dependent prostate cancer cell line LNCaP and androgen-independent prostate cancer cell line DU145 were analyzed by heavy methyl stable isotope labeling with amino acids in cell culture (SILAC) coupled with 2D LC-MS/MS. Western blotting was used to verify the results from MS. The differential expression of related methylases and demethylases was tested by real-time PCR. RESULTS：Five methylation sites on histone H3 were found in both cell lines, the patterns of which were as follows: H3K14me2, H3R17me1, H3K36me1, H3K36me2, H3K36me3, H3R72me2, H3K79me1 and H3K79me2. There were 2 different peptides both containing methylated H3K36, “KSAPATGGVKKPHR” and “KSAPSTGGVKKPHR”, which were different from the 31th amino acid residue “A” and “S”. The former peptide belonging to histone H3 variants, H31T, H31 and H32, was mainly identified in DU145 cells, the total peptide counts of which were much more than that of the latter peptide belonging to histone H3 variant H31T, suggesting that these 2 cell lines expressed different histone H3 variants. Mono- and dimethylation of H3K36 were not different between these 2 cell lines, but the trimethylation was significantly higher in DU145 cells than that in LNCaP cells. Many H3K36 demethyltransferases were decreased in DU145 cells compared with LNCaP cells. CONCLUSION：The differential expression of histone H3 variants and H3K36 demethyltransferases may result in up-regulation of H3K36 tri-methylation during the evolution of prostate cancer from an androgen-dependent state to an androgen-independent state.