Focal adhesion kinase regulates the activity of the osmosensitive transcription factor TonEBP/NFAT5 under hypertonic conditions.
TonEBP/NFAT5 is a major regulator of the urinary concentrating process and is essential for the osmoadaptation of renal medullary cells. Focal adhesion kinase (FAK) is a mechanosensitive non-receptor protein tyrosine kinase expressed abundantly in the renal medulla. Since osmotic stress causes cell shrinkage, the present study investigated the contribution of FAK on TonEBP/NFAT5 activation. Osmotic stress induced time-dependent activation of FAK as evidenced by phosphorylation at Tyr-397, and furosemide reduces FAK Tyr-397 phosphorylation in the rat renal medulla. Both pharmacological inhibition of FAK and siRNA-mediated knockdown of FAK drastically reduced TonEBP/NFAT5 transcriptional activity and target gene expression in HEK293 cells. This effect was not mediated by impaired nuclear translocation or by reduced transactivating activity of TonEBP/NFAT5. However, TonEBP/NFAT5 abundance under hypertonic conditions was diminished by 50% by FAK inhibition or siRNA knockdown of FAK. FAK inhibition only marginally reduced transcription of the TonEBP/NFAT5 gene. Rather, TonEBP/NFAT5 mRNA stability was diminished significantly by FAK inhibition, which correlated with reduced reporter activity of the TonEBP/NFAT5 mRNA 3' untranslated region (3'-UTR). In conclusion, FAK is a major regulator of TonEBP/NFAT5 activity by increasing its abundance via stabilization of the mRNA. This in turn, depends on the presence of the TonEBP/NFAT5 3'-UTR.
Division of Nephrology, Medical Clinic and Policlinic IV, University of Munich Munich, Germany ; Department of Cellular Physiology, University of Munich Munich, Germany.
Wolfgang
W
Neuhofer
Department of Cellular Physiology, University of Munich Munich, Germany.
Christoph
C
Küper
Division of Nephrology, Medical Clinic and Policlinic IV, University of Munich Munich, Germany.
Julia
J
Lichtnekert
Department of Radiology, Klinikum Rechts der Isar, Technical University of Munich Munich, Germany.
Konstantin
K
Holzapfel
Division of Nephrology, Medical Clinic and Policlinic IV, University of Munich Munich, Germany.
Khader V
KV
Rupanagudi
Department of Cellular Physiology, University of Munich Munich, Germany.
Maria-Luisa
ML
Fraek
Center of Anatomy, Institute of Functional and Applied Anatomy, Hannover Medical School Hannover, Germany.
Helmut
H
Bartels
Department of Cellular Physiology, University of Munich Munich, Germany.
Franz-Xaver
FX
Beck
TonEBP/NFAT5; focal adhesion kinase; gene regulation; osmotic stress; renal medulla