Cytokine-Induced Slowing of STAT3 Nuclear Import; Faster Basal Trafficking of the STAT3[beta] Isoform

The STAT3 signal transducer and activator of transcription is a key mediator of gene transcription in response to cytokines such as oncostatin M (OSM). We performed direct live cell imaging of GFP-tagged STAT3 proteins for the first time, showing transient relocalization of STAT3[alpha] to the nucleus following OSM exposure, in contrast to sustained nuclear relocalization of the shorter STAT3[beta] spliceform. To explore this further, we applied fluorescence recovery after photobleaching (FRAP) to determine the nuclear import kinetics of STAT3[alpha] and [beta], as well as of a C-terminal truncation derivative STAT3[Delta]C comprising only the sequence shared by the spliceforms, in the absence or presence of OSM. The rates of basal nuclear import for STAT3[beta] and STAT3[Delta]C were significantly faster than those for STAT3[alpha]. Strikingly, OSM slowed the import rates of all the three STAT3 proteins, whereas the import rates of GFP alone or a classical importin-mediated cargo were unaffected, with analysis of Y705F mutant derivatives for all the three STAT3 constructs, or of a S727A mutant within the unique C-terminus of STAT3[alpha], reinforcing the contribution of specific phosphorylation to the cytokine-stimulated changes. The results introduce a new paradigm where cytokine treatment prolongs nuclear retention simultaneous with decreasing rather than increasing the rate of nuclear import. [PUBLICATION ABSTRACT]