Beta-subunit of cardiac Na+-K+-ATPase dictates the concentration of the functional enzyme in caveolae |
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Authors: | Liu Lijun Askari Amir |
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Institution: | Department of Physiology, Pharmacology, Metabolism, and Cardiovascular Sciences, Medical Univ. of Ohio, 3035 Arlington Ave., Toledo, OH 43614-5804, USA. |
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Abstract: | Previous studies showed the presence of a significant fraction of Na+-K+-ATPase -subunits in cardiac myocyte caveolae, suggesting the caveolar interactions of Na+-K+-ATPase with its signaling partners. Because both - and -subunits are required for ATPase activity, to clarify the status of the pumping function of caveolar Na+-K+-ATPase, we have examined the relative distribution of two major subunit isoforms ( 1 and 1) in caveolar and noncaveolar membranes of adult rat cardiac myocytes. When cell lysates treated with high salt (Na2CO3 or KCl) concentrations were fractionated by a standard density gradient procedure, the resulting light caveolar membranes contained 3040% of 1-subunits and 8090% of 1-subunits. Use of Na2CO3 was shown to inactivate Na+-K+-ATPase; however, caveolar membranes obtained by the KCl procedure were not denatured and contained 75% of total myocyte Na+-K+-ATPase activity. Sealed isolated caveolae exhibited active Na+ transport. Confocal microscopy supported the presence of , -subunits in caveolae, and immunoprecipitation showed the association of the subunits with caveolin oligomers. The findings indicate that cardiac caveolar inpocketings are the primary portals for active Na+-K+ fluxes, and the sites where the pumping and signaling functions of Na+-K+-ATPase are integrated. Preferential concentration of 1-subunit in caveolae was cell specific; it was also noted in neonatal cardiac myocytes but not in fibroblasts and A7r5 cells. Uneven distributions of 1 and 1 in early and late endosomes of myocytes suggested different internalization routes of two subunits as a source of selective localization of active Na+-K+-ATPase in cardiac caveolae. cardiac myocyte; caveolin; oligomer; ouabain; sodium pump |
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