Dynamic analysis of STAT6 signalling in living cells

Abstract

Functional activity of N- and C-terminal fluorescent fusion proteins between STAT6 and EGFP was demonstrated through IL-4-dependent transcriptional activation and nuclear translocation. The N-terminal (EGFP–STAT6) fusion protein appeared to be more active than the C-terminal fusion. In HEK-293 cells both fusion proteins formed fluorescent nuclear foci following IL-4 stimulation, but in HeLa cells nuclear accumulation was homogeneous. Stimulation of the NF-κB pathway through TNFα treatment, or expression of p65–EGFP fusion protein, repressed both basal STAT6-dependent transcriptional activity and the extent of activation in response to IL-4. This indicates a novel mechanism of inhibition of STAT6 signalling by NF-κB activation.