Adenosine blocks IFN-gamma-induced phosphorylation of STAT1 on serine 727 to reduce macrophage activation.
Macrophages are activated by IFN-gamma, a proinflammatory and proatherogenic cytokine that mediates its downstream effects primarily through STAT1. IFN-gamma signaling induces phosphorylation of two STAT1 residues: Tyr(701) (Y701), which facilitates dimerization, nuclear translocation, and DNA binding; and Ser(727) (S727), which enables maximal STAT1 transcription activity. Immunosuppressive molecules such as adenosine in the cellular microenvironment can reduce macrophage inflammatory and atherogenic functions through receptor-mediated signaling pathways. We hypothesized that adenosine achieves these protective effects by interrupting IFN-gamma signaling in activated macrophages. This investigation demonstrates that adding adenosine to IFN-gamma-stimulated murine RAW 264.7 and human THP-1 macrophages results in unique modulation of STAT1 serine and tyrosine phosphorylation events. We show that adenosine inhibits IFN-gamma-induced STAT1 S727 phosphorylation by >30% and phosphoserine-mediated transcriptional activity by 58% but has no effect on phosphorylation of Y701 or receptor-associated JAK tyrosine kinases. Inhibition of the adenosine A(3) receptor with a subtype-specific antagonist (MRS 1191 in RAW 264.7 cells and MRS 1220 in THP-1 cells) reverses this adenosine suppressive effect on STAT1 phosphoserine status by 25-50%. Further, RAW 264.7 A(3) receptor stimulation with Cl-IB-MECA reduces IFN-gamma-induced STAT1 transcriptional activity by 45% and STAT1-dependent gene expression by up to 80%. These data suggest that A(3) receptor signaling is key to adenosine-mediated STAT1 modulation and anti-inflammatory action in IFN-gamma-activated mouse and human macrophages. Because STAT1 plays a key role in IFN-gamma-induced inflammation and foam cell transformation, a better understanding of the mechanisms underlying STAT1 deactivation by adenosine may improve preventative and therapeutic approaches to vascular disease.
Barnholt
Kimberly E
KE
Department of Internal Medicine, Division of Endocrinology, Clinical Nutrition, and Vascular Medicine, University of California, Davis, CA 95616, USA. kebarnholt@ucdavis.edu
Kota
Rama S
RS
Aung
Hnin Hnin
HH
Rutledge
John C
JC
D000241:Adenosine / Q000494:pharmacology*; D058918:Adenosine A3 Receptor Antagonists; D000700:Analgesics / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D004095:Dihydropyridines / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects / Q000276:immunology; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology / Q000494:pharmacology; D053612:Janus Kinases / Q000276:immunology / Q000378:metabolism; D008262:Macrophage Activation / Q000187:drug effects / Q000276:immunology; D008264:Macrophages / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D046228:Microarray Analysis; D010766:Phosphorylation / Q000187:drug effects* / Q000276:immunology; D011799:Quinazolines / Q000494:pharmacology; D043684:Receptor, Adenosine A3 / Q000276:immunology / Q000378:metabolism; D050794:STAT1 Transcription Factor / Q000037:antagonists & inhibitors / Q000276:immunology* / Q000378:metabolism; D012694:Serine / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000276:immunology; D015533:Transcriptional Activation / Q000187:drug effects / Q000276:immunology; D014230:Triazoles / Q000494:pharmacology; D014443:Tyrosine / Q000276:immunology / Q000378:metabolism
C110041:9-chloro-2-(2-furyl)-5-phenylacetylamino(1,2,4)triazolo(1,5-c)quinazoline; D058918:Adenosine A3 Receptor Antagonists; D000700:Analgesics; D004095:Dihydropyridines; C107728:MRS 1191; D011799:Quinazolines; D043684:Receptor, Adenosine A3; D050794:STAT1 Transcription Factor; D014230:Triazoles; D014443:Tyrosine; D012694:Serine; D007371:Interferon-gamma; D053612:Janus Kinases; D000241:Adenosine