Dynamics of the STAT3 Transcription Factor: Nuclear Import Dependent on Ran and Importin-β1

The signal transducer and activator of transcription-3 (STAT3) induces transcription of genes that control differentiation, inflammation, proliferation, and tumor cell invasion. Cytokines such as interleukin-6 and interferon stimulate the specific tyrosine phosphorylation of STAT3, which confers its ability to bind consensus DNA targets. In addition, unphosphorylated STAT3 has been demonstrated to induce specific gene expression. STAT3 must gain entrance to the nucleus to impact transcription, however access to the nucleus is a tightly regulated process. Because nuclear trafficking is critical to the function of STAT3, we investigated the molecular mechanisms by which STAT3 is imported to the nucleus. Live cell imaging techniques were used with STAT3 tagged with green fluorescence protein (GFP) or photoactivatable GFP to follow the cellular dynamics of both unphosphorylated and tyrosine phosphorylated forms. Cytokine activation did not alter the rate of STAT3 nuclear import or nuclear export. In addition, Forster resonance energy transfer experiments revealed homomeric interaction of unphosphorylated STAT3 dependent on its amino terminus, but this dimerization is not necessary for its nuclear import. Previous work demonstrated the adapter importin-alpha 3 binds to STAT3 and is required for nuclear import. To determine whether STAT3 nuclear import is mediated by the importin-alpha/importin-beta 1 heterodimer, the effects of siRNA to importin-beta 1 were evaluated. Results indicate STAT3 nuclear import is dependent on the function of importin-beta 1. Since the Ran GTPase is necessary to bind importin-beta 1 in the nucleus for release of importin-alpha-cargo, the effect of a GTPase deficient mutant of Ran was tested. Expression of the Ran interfering mutant inhibited STAT3 nuclear import. This study defines importin-alpha/importin-beta 1/Ran as the molecular mechanism by which STAT3 traffics to the nucleus.