Stress-induced phosphorylation of STAT1 at Ser727 requires p38 mitogen-activated protein kinase whereas IFN-gamma uses a different signaling pathway

STAT1 is an essential transcription factor for macrophage activation by IFN -gamma and requires phosphorylation of the C-terminal Ser727 for transcript ional activity. In macrophages, Ser727 phosphorylation in response to bacte rial lipopolysaccharide (LPS), UV irradiation, or TNF-alpha occurred throug h a signaling path sensitive to the p38 mitogen-activated protein kinase (p 38 MAPK) inhibitor SB203580 whereas IFN-gamma-mediated Ser727 phosphorylati on was not inhibited by the drug. Consistently, SB203580 did not affect IFN -gamma-mediated. Stat1-dependent transcription but inhibited its enhancemen t by LPS. Furthermore. LPS, UV irradiation, and TNF-alpha caused activation of p38 MAPK whereas IFN-gamma did not. An essential role for p38 MAPK acti vity in STAT1 Ser727 phosphorylation was confirmed by using cells expressin g an SB203580-resistant p38 MAPK. In such cells, STAT1 Ser727 phosphorylati on in response to UV irradiation was found to be SB203580 insensitive. Targ eted disruption of the mapkap-k2 gene, encoding a kinase downstream of p38 MAPK with a key role in LPS-stimulated TNF-cu production and stress-induced heat shock protein 25 phosphorylation, was without a significant effect on UV-mediated Ser727 phosphorylation. The recombinant Stat1 C terminus was p hosphorylated in vitro by p38MAPK alpha and beta but not by MAPK-activated protein kinase 2. Janus kinase 2 activity, previously reported to be requir ed for IFN-gamma-mediated Ser727 phosphorylation, was not needed for LPS-me diated Ser727 phosphorylation, and activation of Janus kinase 2 did not cau se the appearance of STAT1 Ser727 kinase activity. Our data suggest that ST AT1 is phosphorylated at Ser727 by a stress-activated signaling pathway eit her through p38 MAPK directly or through an unidentified kinase downstream of p38MAPK.