Analysis of the antagonist effect of IFN-alpha on IFN-gamma-induced interferon consensus sequence binding protein...

Interferon consensus sequence binding protein (ICSBP), a member of the interferon regulatory factor family of DNA binding proteins, was recently demonstrated in ICSBP knockout mice to play a critical role in hematopoiesis and virus susceptibility. In macrophages, ICSBP mRNA and protein are strongly induced by IFN-gamma, but only marginally by IFN-alpha/beta. When present concurrently, IFN-alpha/beta antagonizes IFN-gamma-induced ICSBP mRNA and protein synthesis. The unknown mechanism(s) underlying this antagonism may involve competitive interactions between these two IFN species or between molecules of their respective signaling cascades. The data presented demonstrate that IFN-alpha does not interfere with initiation of transmembrane signaling by IFN-gamma and that inhibition of ICSBP mRNA expression by IFN-alpha is independent of new protein synthesis. Nuclear proteins from IFN-gamma-treated or from IFN-alpha plus IFN-gamma-treated cells showed identical binding patterns in EMSAs using a palindromic interferon response element (pIRE) from the ICSBP promoter. These proteins were primarily reactive with antibodies directed against STAT1 alpha and, to a lesser extent, against STAT2 and ISGF3 gamma. However, when a second, upstream IRE-like sequence was evaluated by EMSA, a DNA binding pattern distinct from that seen following exposure to IFN-gamma alone was observed after prolonged stimulation with both IFN-alpha and IFN-gamma. These data suggest a possible novel mechanism for IFN-alpha-induced inhibition of IFN-gamma-induced gene expression.