Nuclear localization of the tyrosine kinase Itk and interaction of its SH3domain with karyopherin alpha (Rch1 alpha)

We report a physical and functional association between the Tec-family tyro sine kinase Itk (Emt/Tsk) and the nuclear import chaperone karyopherin alph a (Rch1 alpha) in human T cells. The Itk-SH3 domain and the Rch1 alpha prol ine-rich (PR) motif were crucial for the Itk/Rch1 alpha constitutive intera ction as demonstrated by directed mutagenesis of the Rch1 alpha PR motif (p roline 242 to alanine, P242A). TCR-CD3 stimulation of Jurkat T cells result ed in increased Itk/Rch1 alpha complex formation, recruitment of karyopheri n beta to the protein complex and Rch1 alpha a tyrosine phosphorylation. An alysis of in vitro kinase reactions with a panel of recombinant glutathione -S-transferase (GST) fusion tyrosine kinases (Itk, Lck, ZAP-70 and Jak3) re vealed that only GST-Itk efficiently phosphorylated a recombinant GST-Rch1 alpha fusion. We observed constitutive nuclear localization of Itk that was up-regulated following either TCR-CD3 stimulation or over-expression of wi ld-type Rch1 alpha in T cells. Further, nuclear localization of Itk and TCR -CD3-mediated IL-2 production were significantly down-regulated following e xpression of the Rch1 alpha -P242A mutant, implicating a role for Rch1 alph a in the nuclear translocation of Itk.