Hyperosmotic Stress Strongly Potentiates Serum Response Factor (SRF)-Dependent Transcriptional Activity in Ehrlich Lettre Ascites Cells Through a Mechanism Involving p38 Mitogen-Activated Protein Kinase

Long-term osmotic stress results in altered gene transcription, however, with the exception of the TonE/TonEBP system, the underlying mechanisms are poorly understood. We previously showed that upon osmotic shrinkage of Ehrlich Lettre ' Ascites (ELA) fibroblasts, the MEK1-ERK1/2 pathway is transiently inhibited while p38 MAPK is activated, in turn impacting on cell survival (Pedersen et al., 2007, Cell Physiol Biochem 20: 735-750). Here, we show that downstream of these kinases, two transcription factors with major roles in control of cell proliferation and death, serum response factor (SRF) and cAMP response element-binding protein (CREB) are differentially regulated in ELA cells. SRF Ser 103 phosphorylation and SRF-dependent transcriptional activity were strongly augmented 5-30 min and 24 h, respectively, after hyperosmotic stress (50% increase in extracellular ionic strength), in a p38 MAPK-dependent manner. In contrast, CREB Ser 133 was transiently dephosphorylated upon osmotic shrinkage. The ERK1/2 effector ribosomal S kinase (RSK) and the ERK1/2and p38 MAPK effector mitogen-stress-activated protein kinase 1 (MSK1) both phosphorylate CREB at Ser 133. RSK and MSK1 were dephosphorylated within 5 min of shrinkage. MSK1 phosphorylation recovered within 30 min in a p38-MAPK-dependent manner. CREB was transiently dephosphorylated after shrinkage in a manner exacerbated by p38 MAPK inhibition or MSK1 knockdown, but unaffected by inhibition of RSK. In conclusion, in ELA cells, hyperosmotic stress activates SRF in a p38 MAPK-dependent manner and transiently inactivates CREB, likely due to MSK1 inactivation. We suggest that these events contribute to shrinkage-induced changes in gene transcription and death/survival balance. J. Cell. Physiol. 226: 2857-2868, 2011. (C) 2011 Wiley-Liss, Inc.