STAT3 regulation by S-nitrosylation: implication for inflammatory disease.
OBJECTIVE
S-nitrosylation and S-glutathionylation, redox-based modifications of protein thiols, are recently emerging as important signaling mechanisms. In this study, we assessed S-nitrosylation-based regulation of Janus-activated kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) pathway that plays critical roles in immune/inflammatory responses and tumorigenesis.


RESULTS
Our studies show that STAT3 in stimulated microglia underwent two distinct redox-dependent modifications, S-nitrosylation and S-glutathionylation. STAT3 S-nitrosylation was associated with inducible nitric oxide synthase (iNOS)-produced nitric oxide (NO) and S-nitrosoglutathione (GSNO), whereas S-glutathionylation of STAT3 was associated with cellular oxidative stress. NO produced by iNOS or treatment of microglia with exogenous GSNO inhibited STAT3 activation via inhibiting STAT3 phosphorylation (Tyr(705)). Consequently, the interleukin-6 (IL-6)-induced microglial proliferation and associated gene expressions were also reduced. In cell-free kinase assay using purified JAK2 and STAT3, STAT3 phosphorylation was inhibited by its selective preincubation with GSNO, but not by preincubation of JAK2 with GSNO, indicating that GSNO-mediated mechanisms inhibit STAT3 phosphorylation through S-nitrosylation of STAT3 rather than JAK2. In this study, we identified that Cys(259) was the target Cys residue of GSNO-mediated S-nitrosylation of STAT3. The replacement of Cys(259) residue with Ala abolished the inhibitory role of GSNO in IL-6-induced STAT3 phosphorylation and transactivation, suggesting the role of Cys(259) S-nitrosylation in STAT3 phosphorylation.


METHODS
Microglial proliferation is regulated by NO via S-nitrosylation of STAT3 (Cys(259)) and inhibition of STAT3 (Tyr(705)) phosphorylation.


CONCLUSIONS
Our results indicate the regulation of STAT3 by NO-based post-translational modification (S-nitrosylation). These findings have important implications for the development of new therapeutics targeting STAT3 for treating diseases associated with inflammatory/immune responses and abnormal cell proliferation, including cancer.
1 Department of Pediatrics, Medical University of South Carolina , Charleston, South Carolina.
Jinsu
J
Kim
Je-Seong
JS
Won
Avtar K
AK
Singh
Anand K
AK
Sharma
Inderjit
I
Singh
D000818:Animals; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D004194:Disease; D007249:Inflammation / Q000378:metabolism*; D051379:Mice; D009569:Nitric Oxide / Q000378:metabolism* / Q000494:pharmacology; D010084:Oxidation-Reduction; D010766:Phosphorylation / Q000187:drug effects; D050796:STAT3 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
D050796:STAT3 Transcription Factor; C494087:Stat3 protein, mouse; D009569:Nitric Oxide