Fyn and p38 Signaling Are Both Required for Maximal Hypertonic Activation of the Osmotic Response Element-binding Protein/Tonicity-responsive Enhancer-binding Protein (OREBP/TonEBP)

Abstract

When cells are challenged by hyperosmotic stress, one of the crucial adaptive responses is the expression of osmoprotective genes that are responsible for raising the intracellular level of compatible osmolytes such as sorbitol, betaine, and myo-inositol. This is achieved by the activation of the transcription factor called OREBP (also known as TonEBP or NFAT5) that specifically binds to the osmotic response element (ORE) or tonicity-responsive enhancer that enhances the transcription of these genes. Here we show that p38, a subgroup of the mitogen-activated kinases activated by hypertonic stress, and Fyn, a shrinkageactivated tyrosine kinase, are both involved in the hypertonic activation of OREBP/TonEBP. Inhibition of p38 by SB203580 or by the dominant negative p38 mutant partially blocked the hypertonic induction of ORE reporter (reporter gene regulated by ORE). Similarly, hypertonic activation of ORE reporter was partially blocked by pharmacological inhibition of Fyn or by a dominant negative Fyn and was attenuated in Fyn-deficient cells. Importantly, inhibiting p38 in Fyn-deficient cells almost completely abolished the hypertonic induction of ORE reporter activity, indicating that p38 and Fyn are the major signaling pathways for the hypertonic activation of OREBP/TonEBP. Further we show that the transactivation domain of OREBP/TonEBP is the target of p38-and Fyn-mediated hypertonic activation. These results indicate a dual control in regulating the expression of the osmoprotective genes in mammalian cells.

Degradation of NFAT5, a Transcriptional Regulator of Osmotic Stress-related Genes, Is a Critical Event for Doxorubicin-induced Cytotoxicity in Cardiac Myocytes

“…Hyperosmotic stress enhances translocation of NFAT5 into the nucleus through phosphorylation by signal molecules, such as ATM, Fyn, p38, and protein kinase A (32)(33)(34). In this study, we demonstrated, for the first time, that NFAT5 activity was regulated through the proteasome-mediated proteolysis pathway under cytotoxic conditions, proposing a novel regulatory mechanism of NFAT5 activity.…”

“…Taken together, it is likely that Dox-induced degradation of NFAT5 is mediated by a proteasome-mediated proteolytic process in an ubiquitinindependent manner. NFAT5 activity is regulated by some protein kinases (32)(33)(34). NFAT5 is phosphorylated under hypertonic condition, and NFAT5 phosphorylation is shown to be a critical event for nuclear translocation, followed by DNA binding (41,42).…”

Hypertonicity-Induced Expression of Monocyte Chemoattractant Protein-1 through a Novel Cis-Acting Element and MAPK Signaling Pathways

“…As shown in Fig. 10, p38MAPK and MEK-ERK inhibitors, but not the JNK inhibitor, blocked the nuclear translocation of TonEBP/OREBP/NFAT5 induced by NaCl or mannitol, suggesting that p38MAPK and MEK-ERK signaling pathways (44,45) are associated with the nuclear distribution of TonEBP/ OREBP/NFAT5 mediated by hypertonicity. This result is also consistent with the effects of MAPK inhibitors on MCP1 mRNA and protein levels, as shown in Figs.…”

ATM, a DNA damage-inducible kinase, contributes to activation by high NaCl of the transcription factor TonEBP/OREBP

“…These include PKAc (11), p38 (9, 10), and Fyn (10). Like ATM, each of them is necessary for full activation of TonEBP͞OREBP transcriptional activity by high NaCl, but inhibition of any of them, singly, does not fully prevent the activation.…”

“…There is evidence that several protein kinases participate in activation of TonEBP͞OREBP by hypertonicity, including p38 (9, 10), Fyn (10), and protein kinase A (PKAc) (11). However, there might be additional ones.…”