Figure 1
From: Structure-function analysis indicates that sumoylation modulates DNA-binding activity of STAT1
Sumoylated STAT1 is target for SENP1-mediated desumoylation. (A) SENP1 desumoylates STAT1 in Cos-7 cells. Cos-7 cells were transiently transfected with expression vectors containing STAT1-HA, SUMO-1-His and Flag-tagged SENP1 or SENP1 C603S mutant as indicated. After 36 hours the cells were lysed and total cell lysates were immunoprecipitated with anti-STAT1 antibody and immunoblotted with anti-SUMO-1 and anti-STAT1 antibodies. SENP1 protein levels were verified by immunoblotting with anti-Flag antibody. (B) SENP1 overexpression augments STAT1 transcriptional activity in cultured cells. HeLa cells were transfected with either empty vector or with increasing amount of Flag-tagged WT SENP1 or SENP1 C603S together with GAS-luciferase reporter and CMV-β-galactosidase reporter for normalizing transfection efficiency. After 36 hours cells were starved over night and stimulated with 100 ng/ml of human IFN-γ for 6 hours prior to cell lysis. Luciferase values were normalized against β-galactosidase values. SENP1 and endogenous STAT1 expression levels were verified from the lysates by immunoblotting using anti-Flag and anti-STAT1 antibodies, respectively. Error bars indicate SD from 3 independent transfections. IB: immunoblot, IP: immunoprecipitation.