NDLI: Protein Kinase-mediated Regulation of the Na+/H+ Exchanger in the Rat Myocardium by Mitogen-activated Protein Kinase-dependent Pathways

Content Provider	Journal of Biological Chemistry (JBC)
Author	Moor, Andrea N. Fliegel, Larry
Abstract	We examined regulation of the Na+/H+ exchanger isoform 1 by phosphorylation in the rat myocardium. We utilized cell extracts from adult rat hearts, adult rat extracts fractionated by fast performance liquid chromatography, and extracts from cultured neonatal cardiac myocytes. The carboxyl-terminal 178 amino acids of the Na+/H+ exchanger were expressed inEscherichia coli fused with glutathioneS-transferase. The purified protein was used as a substrate for in vitro phosphorylation and in-gel kinase assays. Unfractionated extracts from neonatal myocytes or adult hearts phosphorylated the COOH-terminal domain of the antiporter. Western blot analysis revealed that mitogen-activated protein (MAP) kinase (44 and 42 kDa) and p90rsk (90 kDa) were present in specific fractions of cardiac extracts that phosphorylated the COOH-terminal protein. In-gel kinase assays confirmed that protein kinases of approximately 44 and 90 kDa could phosphorylate this domain. MAP kinase and p90rsk-dependent phosphorylation of the antiporter could be demonstrated by immunoprecipitation of these kinases from extracts of neonatal cardiac myocytes. PD98059, a mitogen-activated protein kinase kinase inhibitor, decreased MAP kinase and p90rsk phosphorylation of the antiporter and abolished serum and endothelin 1-stimulated increases in steady-state pHi. These results confirm the presence of MAP kinase-dependent phosphorylation in the regulation of the Na+/H+exchanger in the rat myocardium and suggest an important role for p90rsk phosphorylation in regulation of the protein by endothelin-mediated stimulation of the antiporter.
Related Links	http://www.jbc.org/content/274/33/22985.abstract
Ending Page	22992
Starting Page	22985
Page Count	8
File Format	HTM / HTML PDF
ISSN	00219258
Journal	Journal of Biological Chemistry (JBC)
Issue Number	33
Volume Number	274
DOI	10.1074/jbc.274.33.22985
e-ISSN	1083351X
Language	English
Publisher	American Society for Biochemistry and Molecular Biology
Publisher Date	1999-08-13
Access Restriction	Open
Subject Keyword	Na+/H+ exchanger isoform-1 (NHE1) Extracellular signal regulated kinase (ERK) Fast performance liquid chromatography (FPLC) Glutathione S-transferase (GST) Mitogen-activated protein (MAP) Mitogen-activated protein kinase kinase (MEK) Intracellular pH (pHi) 4-morpholinepropanesulfonic acid (MOPS) Polyacrylamide gel electrophoresis (PAGE) Fetal bovine serum (FBS) Radioimmune precipitation buffer (RIPA) Endothelin (ET) MEMBRANES AND BIOENERGETICS
Content Type	Text
Resource Type	Article
Subject	Cell Biology Biochemistry Molecular Biology

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