BETA-INTERFERON AND ONCOSTATIN-M ACTIVATE RAF-1 AND MITOGEN-ACTIVATEDPROTEIN-KINASE THROUGH A JAK1-DEPENDENT PATHWAY

Activation of early response genes by interferons (IFNs) and other cyt okines requires tyrosine phosphorylation of a family of transcription factors termed signal transducers and activators of transcription (Sta ts), The Janus family of tyrosine kinases (Jak1, Jak2, Jak3, and Tyk2) is required for cytokine-induced tyrosine phosphorylation and dimeriz ation of the Stat proteins, In order for IFNs to stimulate maximal exp ression of Stat1 alpha-regulated genes, phosphorylation of a serine re sidue in the carboxy terminus by mitogen-activated protein kinase (MAP K) is also required, In HeLa cells, both IFN-beta and oncostatin M (OS M) stimulated MAPK and Raf-l enzyme activity, in addition to Stat1 and Stat3 tyrosine phosphorylation, OSM stimulation of Raf-l correlated w ith GTP loading of Ras, whereas IFN-beta activation of Raf-l was Ras i ndependent, IFN-beta- and OSM-induced Raf-l activity could be coimmuno precipitated with either Jak1 or Tyk2. Furthermore, HeLa cells lacking Jak1 displayed no activation of STAT1 alpha, STAT3, and Raf-l by IFN- beta or OSM and also demonstrated no increase in the relative level of GTP-bound p21(ras) in response to OSM, The requirement for Jak1 for I FN-beta- and OSM-induced activation of Raf-l was also seen in Jak1-def icient U4A fibrosarcoma cells, Interestingly, basal MAPK, but not Raf- l, activity was constitutively enhanced in Jak1-deficient HeLa cells, Transient expression of Jak1 in both Jak-deficient HeLa cells and U4A cells reconstituted the ability of IFN-beta and OSM to activate Raf-l and decreased the basal activity of MAPK, while expression of a kinase -inactive form of the protein showed no effect. Moreover, U4A cells se lected for stable expression of Jak1, or COS cells transiently express ing Jak1 or Tyk2 but not Jak3, exhibited enhanced Raf-l activity, Ther efore, it appears that Jak1 is required for Raf-l activation by both I FN-beta and OSM. These results provide evidence for a link between the Jaks and the Raf/MAPK signaling pathways.