Fig 1: Ribosomes have a SetDB1-dependent H3K9 methyltransferase activity. (A) Increasing amounts of ribosomes (0.2 and 0.6 ug of RNA) were analyzed by Western blot, as indicated. (B) Western blot analysis of samples derived from immunoprecipitation assays using 50 ug of ribosomal RNA with antibodies against SetDB1 and RPS3a, as well as rabbit antibodies, as negative control. (C) Western blot of HeLa cell extracts treated with 30 nM of either control or SetDB1 siRNA for 72 h, blotted as indicated. (D) Western blot of ribosomes purified by sucrose gradient from HeLa cells treated with 30 nM of either control or SetDB1 siRNA for 72 h, blotted as indicated. 50 ug of S100 cytosolic extract was loaded as a positive control. (E) Histone methyltransferase assay using 3H-SAM, 5 μM of histone H3 peptides from amino acids 1–20 that were either unmodified or trimethylated at the lysine 9, and ribosomes purified by sucrose gradient from HeLa cells treated with 30 nM of either control or SetDB1 siRNA for 72 h. H3K9 methylation value was obtained subtracting the cpms obtained in the reaction using H3(1–20)K9me3 peptide from the cpms obtained in the reaction containing the H3(1–20) peptide. Standard deviation was obtained from triplicates.