Fig. 4
From: SUMO3 modification by PIAS1 modulates androgen receptor cellular distribution and stability
PIAS1 itself was modified by SUMO3 at 117th lysine residue. (a) DU145 cells were cotransfected with plasmids as indicated for 48 h. Whole-cell lysates were immunoprepapited with anti-AR antibody and then analyzed by immunoblot analysis using the indicated antibodies against AR (IP, top panel) and PIAS1 (IB, second panel). Whole-cell lysates (Input) were also immunoblotted with anti-AR (third panel), anti-PIAS1 (fourth panel) or anti-actin (bottom panel) antibodies. (b) DU145 cells were co-transfected with plasmids as indicated for different transfection periods (24 h, 48 h, 72 h and 96 h). Whole cell lysates generated together were immunoprepapited with anti-myc antibody. The myc-immunoprepapite was then detected by anti-myc (IP, top panel) and anti-SUMO3 immunoblotting (IB, second panel). Whole-cell lysates (Input) were also immunoblotted with anti-myc (third panel) or anti-actin (bottom panel) antibodies. (c) Diagrammatic representation of putative typical sumoylation site on human PIAS1 sequense predicted by using the GPS-SUMO software. Analysis of human PIAS1 sequence indicated the presence of only one putative typical sumoylation site, Lys-117, which located near to the PINIT domain of PIAS1. (d) DU145 cells were co-transfected with plasmids as indicated for 48 h and Whole cell lysates were immunoprepapited with anti-myc antibody. The myc-immunoprepapite was then detected by anti-myc (IP, top panel) and anti-SUMO3 immunoblotting (IB, second panel)