Selective STAT3-α or -β expression reveals spliceform-specific phosphorylation kinetics, nuclear retention and distinct gene expression outcomes.
Phosphorylation of STAT3 (signal transducer and activator of transcription 3) is critical for its nuclear import and transcriptional activity. Although a shorter STAT3β spliceform was initially described as a negative regulator of STAT3α, gene knockout studies have revealed that both forms play critical roles. We have expressed STAT3α and STAT3β at comparable levels to facilitate a direct comparison of their functional effects, and have shown their different cytokine-stimulated kinetics of phosphorylation and nuclear translocation. Notably, the sustained nuclear translocation and phosphorylation of STAT3β following cytokine exposure contrasted with a transient nuclear translocation and phosphorylation of STAT3α. Importantly, co-expression of the spliceforms revealed that STAT3β enhanced and prolonged the phosphorylation and nuclear retention of STAT3α, but a STAT3β R609L mutant, with a disrupted SH2 (Src homology 2) domain, was not tyrosine phosphorylated following cytokine stimulation and could not cross-regulate STAT3α. The physiological importance of prolonged phosphorylation and nuclear retention was indicated by transcriptome profiling of STAT3(-/-) cells expressing either STAT3α or STAT3β, revealing the complexity of genes that are up- and down-regulated by the STAT3 spliceforms, including a distinct set of STAT3β-specific genes regulated under basal conditions and after cytokine stimulation. These results highlight STAT3β as a significant transcriptional regulator in its own right, with additional actions to cross-regulate STAT3α phosphorylation and nuclear retention after cytokine stimulation.
Department of Biochemistry and Molecular Biology, Bio21 Molecular Science and Biotechnology Institute, University of Melbourne, Victoria 3010, Australia.
Ivan H W
IH
Ng
Dominic C H
DC
Ng
David A
DA
Jans
Marie A
MA
Bogoyevitch
D017398:Alternative Splicing; D019943:Amino Acid Substitution; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D015870:Gene Expression / Q000187:drug effects; D055786:Gene Knockout Techniques; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation / Q000187:drug effects; D050796:STAT3 Transcription Factor / Q000737:chemistry / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism*; D014443:Tyrosine / Q000737:chemistry; D018909:src Homology Domains
D016207:Cytokines; D017931:DNA Primers; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C494087:Stat3 protein, mouse; D014443:Tyrosine