INTERFERON-GAMMA INDUCTION OF THE HUMAN-LEUKOCYTE ANTIGEN-E GENE IS MEDIATED THROUGH BINDING OF A COMPLEX CONTAINING STAT1-ALPHA TO A DISTINCT INTERFERON-GAMMA-RESPONSIVE ELEMENT

Expression of the human major histocompatibility complex (MHC) class I genes has been shown previously to increase at the transcriptional le vel following exposure to interferon-gamma (IFN-gamma). In this report we have examined the molecular mechanisms involved in the IFN gamma-i nduced transcription of the human MHC class I gene, HLA-E. Functional analysis of CAT reporter gene constructs under the control of the HLA- E promoter transfected into U937 cells revealed the presence of a dist inct IFN-gamma-responsive element, termed the interferon response regi on (IRR), that was necessary and sufficient to mediate the response to IFN-gamma. This cis-acting regulatory sequence contains an imperfect inverted repeat; the 5'-half of the IRR resembles the IFN-gamma activa tion site (GAS), and the 3'-half of the IRR resembles the interferon-s timulated response element (ISRE). Gel mobility shift assays demonstra ted that the IRR bound a single, specific, IFN-gamma-induced complex ( IRR-AC), which was formed rapidly following treatment with IFN-gamma a nd was independent of protein synthesis. Competition experiments with GAS and ISRE sequences from other IFN-inducible genes showed that GAS sequences competed for the IRR-AC, whereas ISRE sequences did not comp ete. Mutational analysis demonstrated that point mutations in either t he 5'-half or 3'-half of the IRR prevented binding of the complex and abrogated or markedly reduced the IFN-gamma responsiveness of reporter gene constructs. Supershift analysis revealed that the IRR-AC contain s a factor that was recognized by antibodies specific for the protein STAT1 alpha (signal transducer and activator of transcription). Taken together, these findings suggest that the mechanism of IFN-gamma-induc ed transcription of the HLA-E gene is distinct from that of other MHC class I genes.