ERK1/2 hijack STAT3 via Thr714 phosphorylation to suppress gene expression.

Hyperactivation of STAT3 has been observed in various tumor types. However, we observed that STAT3 transcriptional activity was undetectable in cells exhibiting high levels of Tyr705 phosphorylation when Thr714 was hyperphosphorylated. The hydroxyl group of Thr714 was indispensable for STAT3 transcriptional activity, and Thr714 mutations were epistatic to other STAT3 modifications. While STAT3-Y705F functioned as a loss-of-function mutant, STAT3-T714E acted as a gain-of-function mutant that repressed transcription. Consistent with these findings, Stat3+/T714E mice exhibited severe primary immunodeficiency, reduced fertility, and increased mortality. Because Thr714 contains an S/T-P motif, it shares upstream kinases with Ser727. Notably, Thr714 phosphorylation exhibited marked regional heterogeneity in lung tumor tissues. Transcriptomic analyses revealed that, in KRAS-driven cancer cells, STAT3 predominantly functions as a transcriptional repressor. Treatment with MAPK inhibitors induced Thr714 dephosphorylation, thereby enhancing feedback STAT3 activation. Collectively, these findings demonstrate that the transcriptional activator STAT3 can be converted into a transcriptional repressor through ERK1/2-mediated phosphorylation of Thr714.