Regulation of Na,K-adenosine triphosphatase gene expression by sodium ions in cultured neonatal rat cardiocytes.
Na,K-ATPase (Na,K-pump) plays an important role in the regulation of intracellular ion composition. The purpose of this study is to determine whether Na+ regulates the levels of mRNA coding for Na,K-ATPase alpha and beta subunits in cultured neonatal rat cardiocytes. We measured intracellular Na+ levels ([Na+]i) in cardiocytes using a Na(+)-sensitive fluorescence dye (SBFI). 1 mM ouabain caused a significant increase in [Na+]i in cardiocytes; from 12.8 +/- 0.3 to 28.8 +/- 1.8 mM. Exposure of cardiocytes to 1 mM ouabain resulted in a three- to fourfold increase in alpha 1, alpha 2, and alpha 3 mRNA accumulation, and an approximate two-fold increase in beta 1 mRNA accumulation. A maximum elevation was reached at 60 min in both cases. The ouabain-induced alpha 1 mRNA accumulation was still observed in the Ca(2+)-free culture medium. Exposure of cardiocytes to 10 microM monensin in the absence of extracellular Ca2+ also resulted in a threefold increase in alpha 1 mRNA accumulation. The increased alpha 1 mRNA expression by 1 mM ouabain was associated with a fourfold increase in alpha 1 subunit protein accumulation. Transfection experiments with chimeric plasmids containing 5'-flanking sequences of alpha 1, alpha 2, and alpha 3 isoform genes and a luciferase reporter gene revealed that 1 mM ouabain caused a twofold increase in luciferase activity in each alpha system. These results suggest that Na+ directly regulates Na,K-ATPase gene expression in cardiocytes. The transfection study further supports the premise that Na(+)-responsive elements are located within the 5'-flanking sequences of each alpha isoform gene.
Yamamoto
K
K
Department of Cardiology, Jichi Medical School, Tochigi, Japan.
Ikeda
U
U
Seino
Y
Y
Tsuruya
Y
Y
Oguchi
A
A
Okada
K
K
Ishikawa
S
S
Saito
T
T
Kawakami
K
K
Hara
Y
Y
D000818:Animals; D000831:Animals, Newborn; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D002478:Cells, Cultured; D004533:Egtazic Acid / Q000494:pharmacology; D004790:Enzyme Induction; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D006321:Heart / Q000187:drug effects; D007527:Isoenzymes / Q000096:biosynthesis; D008156:Luciferases / Q000096:biosynthesis; D046911:Macromolecular Substances; D009206:Myocardium / Q000201:enzymology* / Q000378:metabolism; D010042:Ouabain / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D012964:Sodium / Q000378:metabolism / Q000502:physiology*; D000254:Sodium-Potassium-Exchanging ATPase / Q000096:biosynthesis* / Q000235:genetics
D007527:Isoenzymes; D046911:Macromolecular Substances; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D004533:Egtazic Acid; D010042:Ouabain; D012964:Sodium; D008156:Luciferases; D000254:Sodium-Potassium-Exchanging ATPase; D002118:Calcium