Y705 and S727 are required for the mitochondrial import and transcriptional activities of STAT3, and for regulation of stem cell proliferation.

The STAT3 transcription factor, acting both in the nucleus and mitochondria, maintains embryonic stem cell pluripotency and promotes their proliferation. In this work, using zebrafish, we determined in vivo that mitochondrial STAT3 regulates mtDNA transcription in embryonic and larval stem cell niches and that this activity affects their proliferation rates. As a result, we demonstrated that import of STAT3 inside mitochondria requires Y705 phosphorylation by Jak, whereas its mitochondrial transcriptional activity, as well as its effect on proliferation, depends on the MAPK target S727. These data were confirmed using mouse embryonic stem cells: although the Y705-mutated STAT3 cannot enter mitochondria, the S727 mutation does not affect import into the organelle and is responsible for STAT3-dependent mitochondrial transcription. Surprisingly, STAT3-dependent increase of mitochondrial transcription appears to be independent from STAT3 binding to STAT3-responsive elements. Finally, loss-of-function experiments, with chemical inhibition of the JAK/STAT3 pathway or genetic ablation of stat3 gene, demonstrated that STAT3 is also required for cell proliferation in the intestine of zebrafish.