Stat3 isoforms, α and β, demonstrate distinct intracellular dynamics with prolonged nuclear retention of stat3β mapping to its unique C-terminal end

Two isoforms of Stat3 ( signal transducer and activator of transcription 3) are expressed in cells, alpha(p92) and beta (p83), both derived from a single gene by alternative mRNA splicing. The 55-residue C-terminal transactivation domain of Stat3 alpha is deleted in Stat3 beta and replaced by seven unique C-terminal residues (CT7) whose function remains uncertain. We subcloned the open reading frames of Stat3 alpha and Stat3 beta into the C terminus of green fluorescent protein (GFP). Fluorescent microscopic analysis of HEK293T cells transiently transfected with GFP-Stat3 alpha or GFP-Stat3 beta revealed similar kinetics and cytokine concentration dependence of nuclear accumulation; these findings were confirmed by high throughput microscope analysis of murine embryonic fibroblasts that lacked endogenous Stat3 but stably expressed either GFP-Stat3 alpha or GFP-Stat3 beta. However, although time to half-maximal cytoplasmic reaccumulation after cytokine withdrawal was 15 min for GFP-Stat3 alpha, it was > 180 min for GFP-Stat3 beta. Furthermore, although the intranuclear mobility of GFP-Stat3 alpha was rapid and increased with cytokine stimulation, the intranuclear mobility of GFP-Stat3 beta in unstimulated cells was slower than that of GFP-Stat3 alpha in unstimulated cells and was slowed further following cytokine stimulation. Deletion of the unique CT7 domain from Stat3 beta eliminated prolonged nuclear retention but did not alter its intranuclear mobility. Thus, Stat3 alpha and Stat3 beta have distinct intracellular dynamics, with Stat3 beta exhibiting prolonged nuclear retention and reduced intranuclear mobility especially following ligand stimulation. Prolonged nuclear retention, but not reduced intranuclear mobility, mapped to the CT7 domain of Stat3 beta.