Figure 2.
The AR is methylated by SET9 in vitro and in vivo. (A) An in vitro 3H-SAM methylation assay containing purified SET9 and N- and C-terminal fragments of the AR was incubated for 30 min and samples subject to PAGE, dried and exposed to X-ray film. (B) LNCaP cells grown in steroid-depleted media were treated for 6 h with 10 nM DHT prior to IP using an anti-AR antibody and western analysis using an anti-pan-methyl-lysine antibody (α-pan-methyl-K). (C) LNCaP cells grown in serum-containing media were transiently transfected with either non-silencing (N/S) or SET9 siRNAs for 72 h prior to IP and western analysis as in (B). (D) HEK293T cells were transiently transfected with AR and either SET9 or SET9H297A for 48 h prior to IP and western analysis as in (B).