STAT1 N-terminal domain discriminatively controls type I and type II IFN signaling

The seven signal transducers of transcription (STATs) are cytokine-inducible modular transcription factors. They transmit the stimulation of cells with type I interferons (IFN-alpha/IFN-beta) and type II interferon (IFN-gamma) into altered gene expression patterns. The N-terminal domain (NTD) of STAT1 is a surface for STAT1/STAT1 homodimer and STAT1/STAT2 heterodimer formation and allows the cooperative DNA binding of STAT1. We investigated whether the STAT1 NTD-mediated dimerization affected the IFN-induced tyrosine phosphorylation of STAT1, its nuclear translocation, STAT1-dependent gene expression, and IFN-dependent antiviral defense. We reconstituted human STAT1-negative and STAT2-negative fibrosarcoma cells with STAT1, NTD-mutated STAT1 (STAT1AA), STAT1 with a mutated DNA-binding domain (DBD), or STAT2. We treated these cells with IFN-alpha and IFN-gamma to assess differences between IFN-alpha-induced STAT1 homo- and heterodimers and IFN-gamma-induced STAT1 homodimers. Our data demonstrate that IFNs induce the phosphorylation of STAT1 and STAT1AA at Y701 and their nuclear accumulation. We further reveal that STAT1AA can be phosphorylated in response to IFN-alpha in the absence of STAT2 and that IFN-gamma-induced STAT1AA can activate gene expression directly. However, STAT1AA largely fails to bind STAT2 and to activate IFN-alpha-induced expression of endogenous antiviral STAT1/STAT2 target proteins. Congruent herewith, both an intact STAT1 NTD and STAT2 are indispensable to establish an antiviral state with IFN-alpha. These data provide new insights into the biological importance of the STAT1 NTD.