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Proc Natl Acad Sci U S A. 2002 Dec 24;99(26):16800-5. Epub 2002 Dec 13.

cAMP-independent role of PKA in tonicity-induced transactivation of tonicity-responsive enhancer/ osmotic response element-binding protein.

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  • 1Laboratory of Kidney and Electrolyte Metabolism, National Heart, Lung, and Blood Institute, National Institutes of Health, Department of Health and Human Services, Bethesda, MD 20892, USA.


Hypertonicity-induced increase in activity of the transcription factor tonicity-responsive enhancer/osmotic response element-binding protein (TonEBP/OREBP) protects renal cells by increasing transcription of genes, including those involved in increased accumulation of organic osmolytes. We previously showed that hypertonicity increases transactivating activity of TonEBP/OREBP. Assay with a binary GAL4 transactivation system showed that the 984 C-terminal amino acids of TonEBP/OREBP (amino acids 548-1531) contain a tonicity-dependent transactivation domain (TAD). Also, amino acids 548-1531 undergo tonicity-dependent phosphorylation, and some inhibitors of protein kinases reduce the tonicity-dependent transactivation. In the present studies we examined the role of protein kinase A (PKA).


(i) An inhibitor of PKA (H89) reduces tonicity-dependent increases in transactivation, ORE/TonE reporter activity, and induction of aldose reductase and betaine transporter mRNAs. (ii) Overexpression of the catalytic subunit of PKA (PKAc) increases transactivation activity of amino acids 548-1531 and activity of an ORE/TonE reporter. The increases are much greater under isotonic than under hypertonic conditions. (iii) A dominant-negative PKAc reduces activity of an ORE/TonE reporter. (iv) PKAc activity increases with tonicity but cAMP does not. (v) TonEBP/OREBP and PKAc coimmunoprecipitate. (vi) amino acids 872-1271, including N- and C-terminal polyglutamine stretches, demonstrate tonicity-dependent transactivation, albeit less than amino acids 548-1531, and a similar role for PKA.


(i) PKA plays an important role in TonEBP/OREBP activation of tonicity-dependent gene expression; (ii) PKA activation of TonEBP/OREBP appears to be cAMP-independent; and (iii) amino acids 872-1271 are sufficient for tonicity-dependent transactivation of TonEBP/OREBP.

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