MUTANT-CELL LINES UNRESPONSIVE TO ALPHA BETA AND GAMMA-INTERFERON AREDEFECTIVE IN TYROSINE PHOSPHORYLATION OF ISGF-3-ALPHA COMPONENTS/

The 84-, 91-, and 113-kDa proteins of the ISGF-3 alpha complex are pho sphorylated on tyrosine residues upon alpha interferon (IFN-alpha) tre atment and subsequently translocate to the nucleus together with a 48- kDa subunit. In this study, we investigated the presence and the funct ional status of ISGF-3 alpha subunits and Tyk-2 and JAK1 tyrosine kina ses in mutant HeLa cells defective in the IFN-alpha/beta and -gamma re sponse. Stable cell fusion analysis revealed a single complementation group among one class (class B) of mutants. The class B mutants contai n detectable level of mRNA and proteins of the 84-, 91-, and 113-kDa p roteins, but neither the protein nor mRNA is inducible by IFN-alpha or -gamma. The 91-kDa protein IFN-gamma-activated factor fails to be act ivated into a DNA-binding state after IFN-alpha or -gamma treatment. I n addition, the 91-kDa protein is unable to localize in the nucleus af ter IFN-alpha and -gamma treatment, and the 113-kDa protein fails to t ranslocate after IFN-alpha treatment. Immunoprecipitation studies docu ment a failure of phosphorylation of the 84- or 91-kDa proteins after IPN-alpha or -gamma treatment. Similarly, no tyrosine-phosphorylated 1 13-kDa protein was detected after IFN-alpha treatment. The inability o f class B mutants to phosphorylate the 83-, 91-, or 113-kDa protein on tyrosine residues correlated with the loss of biological response to IFN-alpha and -gamma The genetic defect appears to be the absence of t he tyrosine kinase JAK1. Our data therefore confirm a recent report th at JAK1 plays a critical early signaling role for both IFN-alpha/beta and IFN-gamma systems.