The Expression of Aquaporin-1 in the Medulla of the Kidney Is Dependent on the Transcription Factor Associated with Hypertonicity, TonEBP

Abstract

Expression of aquaporin-1 (AQP1) and -2 (AQP2) channels in the kidney are critical for the maintenance of water homeostasis and the operation of the urinary concentrating mechanism. Hypertonic stress induced in inner medullary (IMCD3) cells by addition of NaCl to the medium substantially up-regulated the mRNA and protein expression of AQP1, suggesting that its activation occurs at a transcriptional and a translational levels. In contrast, no up-regulation of AQP1 was observed when these cells were exposed to the same tonicity by addition of urea. To explore the transcriptional activation of aqp1 under hypertonic stress, we examined the role of the transcription factor associated with hypertonicity, TonEBP. Treatment of IMCD3 cells with the TonEBP inhibitor rottlerin or silencing its expression with specific shRNA technology led to a substantial reduction in AQP1 expression under hypertonic conditions. Moreover, we defined a conserved TonEBP binding site located 811 bp upstream of the aqp1 exon that is essential for its expression. Single site-directed mutation of this TonE site led to a 54 ± 5% (p < 0.01) decrease in AQP1 luciferase-driven activity under hypertonic stress. TonEBP mutant mice display marked decrement in the expression of AQP1 in the inner medulla. In conclusion, these data demonstrate that TonEBP is necessary for the regulation of AQP1 expression in the inner medulla of the kidney under hypertonic conditions.