AN ALTERNATIVE PATHWAY FOR STAT ACTIVATION THAT IS MEDIATED BY THE DIRECT INTERACTION BETWEEN JAK AND STAT

JAK is believed to be an essential tyrosine kinase that mediates signa ls from the cytokine receptor to its downstream events. JAK associates with the cytoplasmic domain of the type I cytokine receptor superfami ly and upon the ligand stimulation it can be activated, resulting in t he receptor phosphorylation. In signaling from gp130, a common signal transducer for the IL-6 family cytokines, STAT3, a transcription facto r that contains an SH2 domain, is recruited by phosphotyrosines on gp1 30 and is subsequently phosphorylated by gp130-associated JAKs. In thi s study, we attempted to find a new target for JAK that is directly ac tivated by JAK, independent of gp130 tyrosine phosphorylation, by usin g a yeast two-hybrid system. Tn the process we found that the JH2 doma in of JAK1, JAK2 or JAK3 could specifically associate with the carboxy -terminal portion of STAT5, but not with STAT3 or STAT1. The interacti on was confirmed using both a transient expression system in a cell li ne and a GST-fusion protein binding assay. Furthermore, we showed that the activation of STAT5 via gp130 did not need any phosphotyrosines o n gp130 while that of STAT3 strictly depended on phosphotyrosines on g p130. Mutations of STAT5 that eliminated the interaction with JAK1 red uced the activation of STAT5 upon the gp130 stimulation, although such mutants could be still activated through erythropoietin receptor. The se results indicate that STAT5 are activated through cytokine receptor s by two distinct mechanisms, one dependent on receptor tyrosine phosp horylation and the other mediated by the JAK-STAT direct interaction.