Using the coiled-coil region of Stat5b as the bait in a yeast two-hybrid screen, we identified the association of Nmi, a protein of unknown function previously reported as an N-Myc interactor. We further show that Nmi interacts with all STATs except Stat2. We evaluated two cytokine systems, IL-2 and IFNgamma, and demonstrate that Nmi augments STAT-mediated transcription in response to these cytokines. Interestingly, Nmi lacks an intrinsic transcriptional activation domain; instead, Nmi enhances the association of CBP/p300 coactivator proteins with Stat1 and Stat5, and together with CBP/p300 can augment IL-2- and IFNgamma-dependent transcription. Therefore, our data not only reveal that Nmi can potentiate STAT-dependent transcription, but also suggest that it can augment coactivator protein recruitment to at least some members of a group of sequence-specific transcription factors.

Interleukin-2 (IL-2) induces heterodimerization of the IL-2 receptor ␤ (IL-2R␤) and ␥ c chains of its receptor and activates the Janus family tyrosine kinases, Jak1 and Jak3. Whereas Jak1 associates with IL-2R␤, Jak3 associates primarily with ␥ c but also with IL-2R␤. We analyzed four IL-2R␤ mutations that diminish IL-2-induced proliferation and found that each also decreased IL-2-induced signal transducer and activator of transcription (STAT) activation. For this reason, and because the mutations were in the IL-2R␤ membrane-proximal region, we investigated and found that each mutation diminished IL-2R␤ association with both Jak1 and Jak3. This suggested that these Jaks might interact with the same region of IL-2R␤; however, certain IL-2R␤ internal deletions and C-terminal truncations differentially affected the association of Jak1 and Jak3. Interestingly, just as Jak1-IL-2R␤ association is Jak3-independent and functionally important, we show that Jak3-IL-2R␤ association is Jak1-independent and implicate this association as being important for IL-2-induced Stat5 activation. Moreover, Jak1 and Jak3 could associate only in the presence of IL-2R␤, suggesting that these kinases can simultaneously bind to IL-2R␤. Thus, our data not only demonstrate that somewhat more distal as well as membrane-proximal cytoplasmic regions of a type I cytokine receptor are important for Jak kinase association but also suggest that two IL-2R␤-Jak kinase interactions are important for IL-2 signaling.

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