MUTATIONAL ANALYSIS OF INTERFERON (IFN) REGULATORY FACTOR-1 AND FACTOR-2 - EFFECTS ON THE INDUCTION OF IFN-BETA GENE-EXPRESSION

Interferon (IFN) regulatory factor 1 (IRF-1) and IRF-2 are structurall y similar but functionally distinct transcription factors that bind to the positive regulatory domains I and III (PRDI/III) within the human IFN-beta promoter. To begin structure-function analysis of IRF-1 and IRF-8, the regulatory potential of carboxyl-terminal deletion mutants was analyzed by co-transfection studies in human cells and was correla ted with DNA binding capacity. Transcriptional repression by IRF-2 was contained within the first 125 amino-terminal amino acids and correla ted directly with IRF-2 DNA binding; deletion to a protein of 100 amin o acids resulted in loss of repression and IRF-2 DNA binding. Thus, th e carboxyl terminus appears dispensible for trans-repression. Hybrid c onstructs which fuse the DNA binding domain of HRF-1 and IRF-2 to the trans-activation domain of NF-kappa B p65 were also generated; both IR F-1/p65 and IRF-2/p65 chimeras were strong transcriptional activators. IRF-2-mediated repression was also dominant over trans-activation by these fusion proteins. The trans-activation region of IRF-1 resides in the carboxyl terminus, primarily carboxyl-terminal to amino acid 250. Mutation of three potential casein kinase II phosphorylation sites wi thin the IRF carboxyl terminus failed to identify an essential site th at contributes to IRF-1 trans-activation potential.