Activated STAT1 suppresses proliferation of cultured rat mesangial cells

Background JAK-STAT signaling has been shown to promote development and pro liferation in lymphopoietic and hematopoietic lineages. We investigated the effect of activated STAT1 on mesangial cell proliferation. Methods. Rat mesangial cells of primary culture (rMCs) were used in the fol lowing experiments: (I) Whole cell lysates were immunoblotted against JAK1 and JAK2. (2) Whole cell lysates and nuclear proteins were extracted from r MCs with or without treatment with interferon-gamma, and immunoblotting was performed against both STAT1 and tyrosine (701)-phosphorylated STAT1. (3) rMCs and rMCs electroporated with either wild-type STAT1, mutated STAT1, or antibody against STAT1 were incubated with interferon-gamma for 20 hours, followed by a further incubation with [H-3]-thymidine for four hours. Results. JAK1, JAK2, and STAT1 were detected in whole cell lysates, suggest ing that JAK-STAT signaling could be activated by interferon-gamma (INF-gam ma). Using an antibody specific for tyrosine-phosphorylated STAT1, we detec ted signal in the INF-gamma-treated nuclear extracts, which showed transloc ation of phosphorylated STAT1 to the nucleus. [H-3]-thymidine incorporation in the presence of INF-gamma was significantly lower than that of control in a dose-dependent manner. The introduction of wild-type STAT1 enhanced th e effect of interferon-gamma and decreased [H-3]-thymidine incorporation, w hereas tyrosine-mutated (Y701F) STAT1 and SH2 domain (R602T)-mutated STAT1 reversed INF-gamma-induced suppression of [H-3]-thymidine incorporation. El ectroinjected antibody against STAT1 increased [H-3]-thymidine incorporatio n upon stimulation with INF-gamma. Conclusion. STAT1 activated by interferon-gamma suppresses mesangial cell p roliferation.