Hypertonicity (most often present as high salinity) is stressful to the cells of virtually all organisms. Cells survive in a hypertonic environment by increasing the transcription of genes whose products catalyze cellular accumulation of compatible osmolytes. In mammals, the kidney medulla is normally hypertonic because of the urinary concentrating mechanism. Cellular accumulation of compatible osmolytes in the renal medulla is catalyzed by the sodium͞ myo-inositol cotransporter (SMIT), the sodium͞chloride͞ betaine cotransporter, and aldose reductase (synthesis of sorbitol). The importance of compatible osmolytes is underscored by the necrotic injury of the renal medulla and subsequent renal failure that results from the inhibition of SMIT in vivo by administration of a specific inhibitor. Tonicity-responsive enhancers (TonE) play a key role in hypertonicity-induced transcriptional stimulation of SMIT, sodium͞ chloride͞betaine cotransporter, and aldose reductase. We report the cDNA cloning of human TonE binding protein (TonEBP), a transcription factor that stimulates transcription through its binding to TonE sequences via a Rel-like DNA binding domain. Western blot and immunohistochemical analyses of cells cultured in hypertonic medium reveal that exposure to hypertonicity elicits slow activation of TonEBP, which is the result of an increase in TonEBP amount and translocation to the nucleus.

Cells in the hypertonic renal medulla maintain their intracellular ion concentration at isotonic levels, despite much higher concentrations of extracellular electrolytes, by accumulating high concentrations of nonperturbing small organic solutes termed osmolytes. Taurine has been identified as a nonperturbing osmolyte in the renal medulla and Madin-Darby canine kidney (MDCK) cells. In hypertonic medium, the increased accumulation of taurine in MDCK cells is the result of increased activity of a Na+-and Cl1-dependent taurine transporter. We have isolated a cDNA encoding a Na+-and Cl-dependent taurine transporter, whose sequence corresponds to a protein of 655 amino acids with igncant amino acid sequence similarity to previously cloned Na+-and Cldependent transporters, incuding the MDCK cell betaine/-aminobutyric acid transporter and several brain neurotransmitter transporters. Northern hybridization indicates that mRNA for the taurine transporter is present in renal cortex and medulla, ileal mucosa, brain, liver, and heart. The abndce of mRNA for the taurine transporter is increased in MDCK cells cultured in hypertonic medium, suggesting that regulation of transport activity by medium hypertonicity occurs at the level of mRNA accumulation. imum velocity (V.,.) of the taurine transporter in the basolateral plasma membrane without change in Km (4).Taurine transport in renal brush border membranes has also been well characterized (5). The activity of the cotransporter in the brush border of the proximal tubule contributes to whole-body homeostasis of taurine; activity increases in animals fed diets deficient in taurine and in sulfur-containing amino acids (6). The addition of 50 pAM taurine to taurine-free medium results in a decrease in the activity of the taurine transporter in MDCK cells (4,7). However, the rate of taurine transport was up-regulated by hypertonicity to the same degree as in cells cultured in taurine-free medium, suggesting that the regulation of taurine transport by hypertonicity and the regulation by medium taurine are independent of each other (4).In this report we describe the cloning of the cDNA for the MDCK cell taurine transporter. The sequence of the cDNAt indicates that the taurine transporter has considerable amino acid sequence similarity to the previously cloned Na+-and Cl--dependent transporters. Northern hybridizations indicate that the abundance of mRNA for the taurine transporter in MDCK cells is regulated by hypertonicity.Taurine (2-aminoethanesulfonic acid) is a major intracellular amino acid in mammals (1). It is involved in a number of important physiological processes, including bile acid conjugation in hepatocytes, modulation of calcium flux and neural excitability, osmoregulation, detoxification, and membrane stabilization (1).The cells of virtually all organisms respond to hypertonicity by the intracellular accumulation ofhigh concentrations of small organic solutes (osmolytes) that, in contrast to high concentrations of electrolytes, do not perturb the function of ...

Tonicity-responsive enhancer binding protein (TonEBP) regulates transcription of tonicity responsive genes such as the sodium-myo-inositol cotransporter (SMIT), the sodium-chloride-betaine cotransporter (BGT1), and aldose reductase (AR). To characterize signals that activate TonEBP in Madin-Darby canine kidney (MDCK) cells, the abundance and nuclear distribution of TonEBP were studied after the osmolality of the culture medium was changed. Hypertonicity but not hyperosmolality is effective in activation of TonEBP as expected. Surprisingly, exposure to hypotonic medium leads to a dramatic downregulation of TonEBP both in abundance and nuclear distribution, indicating that under isotonic conditions, TonEBP is at a low-level activated state and can respond to both increase and decrease in tonicity. Additional experiments suggest that cellular ionic strength is the signal that initiates regulation of TonEBP. The increase in abundance of TonEBP is mediated by an increase in mRNA abundance and a parallel increase in synthesis of TonEBP. The stability of TonEBP mRNA is not affected by hypertonicity indicating that transcription plays a major role in the induction of TonEBP by hypertonicity.

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