MULTIPLE SIGNAL-TRANSDUCTION PATHWAYS LINK NA+ K+-ATPASE TO GROWTH-RELATED GENES IN CARDIAC MYOCYTES - THE ROLES OF RAS AND MITOGEN-ACTIVATED PROTEIN-KINASES/

We showed before that in neonatal rat cardiac myocytes partial inhibit ion of Na+/K+-ATPase by nontoxic concentrations of ouabain causes hype rtrophic growth and transcriptional regulations of genes that are mark ers of cardiac hypertrophy, In view of the suggested roles of Ras and p42/44 mitogen-activated protein kinases (MAPKs) as key mediators of c ardiac hypertrophy, the aim of this work was to explore their roles in ouabain-initiated signal pathways regulating four growth-related gene s of these myocytes, i.e. these for c-Fos, skeletal alpha-actin, atria l natriuretic factor, and the alpha(3)-subunit of Na+/K+-ATPase, Ouaba in caused rapid activations of Ras and p42/44 MAPKs; the latter was su stained longer than 90 min. Using high efficiency adenoviral-mediated expression of a dominant-negative Ras mutant, and a specific inhibitor of MAPK kinase (MEK), activation of Ras-Raf-MEK-p42/44 MAPK cascade b y ouabain was shown. The effects of the mutant Ras, an inhibitor of Ra s farnesylation, and the MEK inhibitor on ouabain-in duced changes in mRNAs of the four genes indicated that (a) skeletal alpha-actin induct ion was dependent on Rat; but not on p42/44 MAPKs, (b) alpha(3) repres sion was depend ent on the Ras-p42/44 MAPK cascade, and (c) induction of c-fos or atrial natriuretic factor gene occurred partly through the Ras-p42/44 MAPK cascade, and partly through pathways independent of R as and p42/44 MAPKs, All ouabain effects required extracellular Ca2+ a nd were attenuated by a Ca2+/calmodulin antagonist or a protein kinase C inhibitor. The findings show that (a) signal pathways linked to sar colemmal Na+/K+-ATPase share early segments involving Ca2+ and protein kinase C, but diverge into multiple branches only some of which invol ve Pas, or p42/44 MAPKs, or both; and (b) there are significant differ ences between this network and the related gene regulatory pathways ac tivated by other hypertrophic stimuli, including those whose responses involve increases in intracellular free Ca2+ through different mechan isms.