SYNERGY BETWEEN INTERFERON-GAMMA AND TUMOR-NECROSIS-FACTOR-ALPHA IN TRANSCRIPTIONAL ACTIVATION IS MEDIATED BY COOPERATION BETWEEN SIGNAL TRANSDUCER AND ACTIVATOR OF TRANSCRIPTION-1 AND NUCLEAR FACTOR KAPPA-B

Interferon-gamma (IFN gamma) and tumor necrosis factor-alpha (TNF alph a) cooperate to induce the expression of many gene products during inf lammation, The present report demonstrates that a portion of this coop erativity is mediated by synergism between two distinct transcription factors: signal transducer and activator of transcription 1 (STAT1) an d nuclear factor kappa B (NF-kappa B). IFN gamma and TNF alpha synergi stically induce expression of mRNAs encoding interferon regulatory fac tor-1 (IRF-1), intercellular adhesion molecule-1, Mig (monokine induce d by gamma-interferon), and RANTES (regulated on activation normal T c ell expressed and secreted) in normal but not STAT1-deficient mouse fi broblasts, indicating a requirement for STAT1, Transient transfection assays in fibroblasts using site-directed mutants of a 1.3-kilobase pa ir sequence of the IRF-1 gene promoter revealed that the synergy was d ependent upon two sequence elements; a STAT binding element and a kapp a B motif. Artificial constructs containing a single copy of both a ST AT binding element and a kappa B motif linked to the herpes virus thym idine kinase promoter were able to mediate synergistic response to IFN gamma and TNF alpha; such response varied with both the relative spac ing and the specific sequence of the regions between these two sites, Cooperatively responsive sequence constructs bound both STAT1 alpha an d NF-kappa B in nuclear extracts prepared from IFN gamma- and/or TNF a lpha-stimulated fibroblasts, although binding of individual factors wa s not cooperative, Thus, the frequently observed synergy between IFN g amma and TNF alpha in promoting inflammatory response depends in part upon cooperation between STAT1 alpha and NF-kappa B, which is most lik ely mediated by their independent interaction with one or more compone nts of the basal transcription complex.