The MKK6-p38 MAPK pathway prolongs the cardiac contractile calcium transient, downregulates SERCA2, and activates NF-AT

Objective: Our goal was to determine if the MKK6-p38 MAPK pathway regulates cardiac intracellular calcium ([Ca2+],). We also tested if MKK6 might influence expression of SERCA2, a calcium regulatory molecule involved in relaxation, and the activity of nuclear factor of activated T-cells (NF-AT), a calcium-regulated transcription factor that participates in pathological responses to pressure-overload. Methods: Neonatal rat ventricular myocytes were transfected with MKK6(Glu), an activator of p38 MAPK. Green fluorescent protein (GFP) was used as transfection marker and [Ca2+](i) was evaluated via indo-1. SERCA2 expression was assayed via Northern and Western techniques. The activity of the rat SERCA2 gene promoter and NF-AT-dependent gene expression were monitored with reporter genes. Myocyte contractility was regulated by electrical pacing. Results: MKK6(Glu) prolonged decay of the contractile calcium transients, downregulated SERCA2 expression, and reduced the activity of the rat SERCA2 gene promoter. Diastolic [Ca2+](i) in myocytes pacing at 1-2 Hz was dramatically increased by MKK6(Glu). NF-AT-dependent gene expression was activated by MKK6(Glu) and by pacing of contractions in a synergistic manner. Overexpression of SERCA2 mitigated the effects of MKK6(Glu) on [Ca2+](i) and NF-AT. Conclusions: The MKK6(Glu)-p38 MAPK pathway prolongs the decay phase of the cardiac contractile calcium by downregulating SERCA2, increasing diastolic [Ca2+](i) which activates NF-AT. The ability of SERCA2 over-expression to reduce NF-AT activity represents a potential novel therapeutic effect of SERCA2 that should be further considered in the development of cardiac gene therapy strategies. (C) 2003 European Society of Cardiology. Published by Elsevier Science B.V. All rights reserved.