Gamma interferon signaling: insights to development of interferon mimetics.

We have developed small peptide mimetics of gamma interferon (IFNgamma), based not on the classical model of IFNgamma initiated signaling by extracellular interaction, but rather on direct intracellular signaling by IFNgamma. IFNgamma, its receptor subunit IFNGR1, and transcription factor STAT1alpha are transported to the nucleus of cells as a complex where IFNgamma provides a classical polycationic nuclear localization sequence (NLS) for such transport. The C terminus of IFNgamma, represented here by the mouse IFNgamma peptide, IFNgamma(95-132), was capable of also forming a complex with IFNGR1 and STAT1alpha when introduced intracellularly and provided the NLS signaling for nuclear transport. Importantly, mouse IFNgamma(95-132) and human IFNgamma(95-134) mimetics both induced an antiviral state and upregulation of MHC class II molecules in cells similar to that of full length IFNgamma. Both IFNgamma and its peptide mimetics bind to an intracellular site, IFNGR1(253-287), on the cytoplasmic domain of receptor subunit IFNGR1. This binding plays a role in tyrosine phosphorylation events, catalyzed by JAK1 and JAK2 kinases that result in the phosphorylation and binding of STAT1alpha to the cytoplasmic domain of IFNGR1. Important structural requirements for IFNgamma mimetic activity are a polycationic NLS and an alpha helix in the mimetics. Finally, chromatin immunoprecipitations and reporter gene studies of IFNgamma and IFNgamma mimetic treated cells indicate that they, along with IFNGR1 and STAT1alpha, bind to the GAS element of IFNgamma activated genes and participate in STAT1alpha transcription. It is important to note that IFNgamma intracellular events played the key role in development of IFNgamma mimetics.