Regulation Of Classical And Non-classical IFN-γ-triggered Signaling

IFNs are important components of the immune system,which generally trigger cellular protective defenses in response to infection.IFN-γ(also known as type Ⅱ IFN)is a paradigm for this,being an important mediator of innate and adaptive immune responses with a key role in clearance of viral and bacterial pathogens.The binding of IFN-y homodimer to IFNGR1 and IFNGR2 results in spatial close of JAK1 and JAK2,leading to phosphorylation of IFNGR1 and JAKs.The phosphorylation of IFNGR1 at Tyr440 by JAKs provides a docking site for the recruitment of the transcription factor signal transducers and activators of transcription 1(STAT1),and the phosphorylated JAK kinases subsequently phosphorylate STAT1 at Tyr701.Phosphorylated STAT1 forms homodimers which tanslocate to the nucleus and bind to the conserved IFN-γ-activated sites(GAS)on the promoters of the IFN-stimulated genes(ISGs)to initiate the transcription of target genes.In addition to the phosphorylation of Tyr701 of STAT1,IFN-γ induces phosphorylation of STAT1 at Ser727,which is mediated by PI3K and the downstream protein kinase C and AKT,and this phosphorylation is critical for the full activation of STAT1.In addition to the well known IFN-γ-JAK-STAT1 pathway,IFN-γ activates additional signal pathways.IKKβ,a master activator of inflammatory response,is required to activate a subset of IFN-γ induced genes.The mechanism and biological function of this non-classical IFN-y signaling are not fully clear.Whether and how additional molecules are involved in the process are not fully clear.In this study,we identified parafibromin as an important component of the IFN-γ-triggered signaling pathways.Overexpression of parafibromin promoted IFN-y-triggered phosphorylation of STAT1 at Tyr701,subsequent expression of downstream genes,and cellular antiviral response,whereas knockdown of parafibromin had opposite effects.Parafibromin interacted with JAK1/2,promoted the interactions of JAK1-JAK2 and JAK1/2-STAT1,and facilitated tyrosine phosphorylation of STAT1 by JAKs after IFN-y stimulation.Our results reveal a previously uncharacterized role of parafibromin in mediating IFN-γ-triggered signaling and cellular effects.In addition,we identified sorting nexin 8(SNX8)as an important component of the non-classical IFN-γ-triggered signaling pathways.Overexpression of SNX8 promoted IFN-γ-triggered phosphorylation of STAT1 at Tyr701,subsequent expresssion of a subset of downstream genes,whereas knockdown or knockout of SNX8 had opposite effects.Snx8-/-mice infected with Listeria monocytogenes exhibited lower cytokine levels and higher bacterial loads in the liver and spleen,resulting in higher lethality.SNX8 interacted with JAK1/IKKβ,and promoted the interaction of JAK1-IKKβ after IFN-γ stimulation.In addition,IFN-γ induced the tyrosine phosphorylation of SNX8 at Tyr95 and Tyr126 by JAKs.The phosphorylated SNX8 recruited IKKβ to JAK1 and subsequently promotes the autophosphorylation of IKKβ.Moreover,JAK1-SNX8-IKKβ complex promoted IFN-γ-triggered phosphorylation of STAT1.Our findings suggest that SNX8 plays an important role in innate immune response against Listeria monocytogenes.In summary,we identified parafibromin and SNX8 as important components of classical and non-classical IFN-γ-triggered signaling pathways.Our study provide a novel mechanism in mediating IFN-γ-triggered signaling and innate immune response.