Abstract
Cardiac plasma membrane Ca2+/Mg2+ ecto-ATPase (myoglein) requires millimolar concentrations of either Ca2+ or Mg2+ for maximal activity. In this paper, we report its localization by employing an antiserum raised against the purified rat cardiac Ca2+/Mg2+ ATPase. As assessed by Western blot analysis, the antiserum and the purified immunoglobulin were specific for Ca2+/Mg2+ ecto-ATPase; no cross reaction was observed towards other membrane bound enzymes such as cardiac sarcoplasmic reticulum Ca2+-pump ATPase or sarcolemmal Ca2+-pump ATPase. On the other hand, the cardiac Ca2+/Mg2+ ecto-ATPase was not recognized by antibodies specific for either cardiac sarcoplasmic reticulum Ca2+-pump ATPase or plasma membrane Ca2+-pump ATPase. Furthermore, the immune serum inhibited the Ca2+/Mg2+ ecto-ATPase activity of the purified enzyme preparation. Immunofluorescence of cardiac tissue sections and neonatal cultured cardiomyocytes with the Ca2+/Mg2+ ecto-ATPase antibodies indicated the localization of Ca2+/Mg2+ ecto-ATPase in association with the plasma membrane of myocytes, in areas of cell-matrix or cell-cell contact. Staining for the Ca2+/Mg2+ ecto-ATPase was not cardiac specific since the antibodies detected the presence of membrane proteins in sections from skeletal muscle, brain, liver and kidney. The results indicate that Ca2+/Mg2+ ecto-ATPase is localized to the plasma membranes of cardiomyocytes as well as other tissues such as brain, liver, kidney and skeletal muscle.
Similar content being viewed by others
References
Dhalla NS, Zhao D: Cell membrane Ca2+/Mg2+ ATPase. Prog Biophys Mol Biol 52: 1–37, 1988
Dhalla NS, Zhao D: Possible role of sarcolemmal Ca2+/Mg2+ ATPase in heart function. Mag Res 2: 161–172, 1989
Plessner L: Ecto-ATPases: Identities and functions. Int Rev Cytol 158: 141–214, 1995
Zhao D, Dhalla NS: Purification and characterization of Ca2+/Mg2+ ATPase from rat heart plasma membrane. Mol Cell Biochem 107: 135–149, 1991
Kannan S, Elimban V, Dakshinamurti K, Dhalla NS: Biophysical characterization of rat cardiac Ca2+/Mg2+ ecto ATPase (Myoglein). Mol Cell Biochem 183: 153–157, 1998
Kannan S, Elimban V, Fandrich RF, Kardami E, Dhalla NS: Evidence that the rat cardiac sarcolemmal Ca2+/Mg2+ ecto ATPase is a cell adhesion molecule. Cardiovasc Pathobiol 1: 1–7, 1997
Malouf NN, Meissner G: Cytochemical localization of a basic ATPase to canine myocardial surface membrane. J Histochem Cytochem 28: 1286–1294, 1980
Slezak J, Okruhlicova L, Tribulova N, Schulze W, Dhalla NS: Renaissance of cytochemical localization of membrane ATPases in the myocardium. Mol Cell Biochem 147: 169–172, 1995
Pitts BJR: Stoichiometry of sodium-calcium exchange in cardiac sarcolemmal vesicles. J Biol Chem 254: 6232–6235, 1979
Ganguly PK, Pierce GN, Dhalla NS: Defective sarcoplasmic reticular calcium transport in diabetic cardiomyopathy. Am J Physiol 244: E528–E535, 1983
Laemmli UK: Cleavage of structural proteins during the assembly of the heart of the bacteriophage T4. Nature 227: 680–689, 1970
Morrissey JH: Silver stain for proteins in polyacrylamide gels: A modified procedure with enhanced uniform sensitivity. Anal Biochem 117: 307–310, 1981
Harlow E, Lane D: Antibodies. A Laboratory Manual. Cold Spring Harbour Laboratory, Cold Spring Harbour, NY, 1983
Kardami E, Fandrich RR: Basic fibroblast growth factor in atria and ventricles of the vertebrate heart. J Cell Biol 109: 1865–1875, 1989
Kardami E, Fandrich RR: Basic fibroblast growth factor in cultured cardiac myocytes. Ann New York Acad Sci 638: 244–255, 1991
Lin SH: Localization of the ecto-ATPase (ecto-nucleotidase) in the rat hepatocyte plasma membrane. Implications for the functions of the ecto-ATPase. J Biol Chem 264: 14403–14407, 1989
Buchweitz O, Bianchi CP: Myocardial magnesium transport: Effect of gramicidin S and epinephrine. Life Sci 55: 1863–1861, 1994
Zhao D, Dhalla NS: Influence of gramicidin S on cardiac membrane Ca2+/Mg2+ ecto-ATPase activities and contractile force development. Can J Physiol Pharmacol 67: 546–552, 1989
Beaudoin AR, Sévigny J, Grondin G, Daoud S, Levesque FP: Purification, characterization, and localization of two ATP di-phosphohydrolase isoforms in bovine heart. Am J Physiol 42: H673–H681, 1997
Lin SH, Russell WE: Two Ca2+-dependent ATPases in rat liver plasma membrane. J Biol Chem 263: 12253–12258, 1988
Cheung PH, Luo W, Qiu Y, Zhang X, Earley K, Millirons P, Lin SH: Structure and function of C-CAM 1. The first immunoglobulin domain is required for intercellular adhesion. J Biol Chem 268: 24303–24310, 1993
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Kannan, S., Elimban, V., Fandrich, R.R. et al. Immunolocalization of the sarcolemmal Ca2+/Mg2+ ecto-ATPase (myoglein) in rat myocardium. Mol Cell Biochem 197, 187–194 (1999). https://doi.org/10.1023/A:1006982708128
Issue Date:
DOI: https://doi.org/10.1023/A:1006982708128