Interferon-gamma signaling in human retinal pigment epithelial cells mediated by STAT1, ICSBP, and IRF-1 transcription factors.

PURPOSE Studies have shown that interferon (IFN)-gamma stimulates expression of intercellular adhesion molecule-1 (ICAM-1), major histocompatibility complex (MHC) class II, interleukin (IL)-6, and inducible nitric oxide synthase and inhibits replication of Toxoplasma gondii in human retinal pigment epithelial (HRPE) cells. The present study was undertaken to investigate the molecular mechanisms of IFN-gamma action. METHODS RNA, whole-cell extracts, and nuclear extracts were prepared from HRPE cells cultured in the presence or absence of IFN-gamma. Activation of IFN-gamma-responsive genes was analyzed by electrophoretic mobility shift assay, reverse transcription-polymerase chain reaction (RT-PCR), western blot analysis, and immunoprecipitation. RESULTS HRPE cells constitutively expressed two members of the IFN regulatory factor (IRF) family of transcription factors, IRF-1 and IRF-2. After exposure to IFN-gamma, transcription of IRF-1 and IFN consensus sequence binding protein (ICSBP) genes were induced; IRF-2 gene transcription was not upregulated. Activation of IFN-gamma-responsive genes was mediated by tyrosine phosphorylation of the signal transducer and activator of transcription (STAT)-1 factor. CONCLUSIONS This study characterized the IFN-gamma signaling pathway in HRPE cells and identified IRF-1, ICSBP, and tyrosine-phosphorylated STAT1 as mediators of IFN-gamma action in these cells. ICSBP is thought to be exclusively used in immunologic responses and has previously been detected only in lymphoid cells. However, the current study shows that ICSBP expression is inducible in HRPE cells, suggesting that it may regulate gene transcription in RPE cells and possibly in other nonimmunologic cell types.