CDK9 regulates hypotonicity‐induced nuclear export of NFAT5

NFAT5, also known as Osmotic Response Element‐binding Protein (OREBP), or Tonicity‐responsive Enhancer‐binding Protein (TonEBP), is an osmo‐ sensitive transcription factor that mediates cellular adaptations to extracellular hypertonic stress. The subcellular localization and activity of NFAT5 are tightly regulated by extracellular tonicity. Earlier we showed that hypotoncity‐induced nuclear export of NFAT5 is regulated by the sequential phosphorylation of two serine residues (ser‐155 and ser‐158), and we also identified Casein Kinase 1 (CK1) as the ser‐158 kinase. In this study, using protein fractionations, in‐vitro kinase assay and mass spectrometric protein identification approaches, we identified Cyclin‐dependent Kinase 9 (CDK9) as a putative ser‐155 kinase. NFAT5 is co‐immunoprecipitated with CDK9 and its regulatory subunit cyclin T. Furthermore, CDK9 inhibitors and siRNAs significantly diminished NFAT5 nuclear export in response to hypotonicity.