Protein Phosphatase 1c Associated with the Cardiac Sodium Calcium Exchanger 1 Regulates Its Activity by Dephosphorylating Serine 68-phosphorylated Phospholemman.

Autor:	Hafver TL; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Hodne K; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway, the Department of Basic Sciences and Aquatic Medicine, Norwegian University of Life Sciences (NMBU), 0454 Oslo, Norway., Wanichawan P; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Aronsen JM; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the Bjørknes College, Oslo, Norway., Dalhus B; the Department of Microbiology, Oslo University Hospital, Rikshospitalet, 0424 Oslo, Norway, the Department of Medical Biochemistry, Institute for Clinical Medicine, University of Oslo, 0424 Oslo, Norway and., Lunde PK; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Lunde M; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Martinsen M; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Enger UH; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Fuller W; the Cardiovascular and Diabetes Medicine, School of Medicine, University of Dundee, Dundee, Scotland, United Kingdom DD1 9SY., Sjaastad I; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Louch WE; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Sejersted OM; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway., Carlson CR; From the Institute for Experimental Medical Research, Oslo University Hospital and University of Oslo, 0450 Oslo, Norway, the KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo, 0316 Oslo, Norway, c.r.carlson@medisin.uio.no.
Jazyk:	angličtina
Zdroj:	The Journal of biological chemistry [J Biol Chem] 2016 Feb 26; Vol. 291 (9), pp. 4561-79. Date of Electronic Publication: 2015 Dec 14.
DOI:	10.1074/jbc.M115.677898
Abstrakt:	The sodium (Na(+))-calcium (Ca(2+)) exchanger 1 (NCX1) is an important regulator of intracellular Ca(2+) homeostasis. Serine 68-phosphorylated phospholemman (pSer-68-PLM) inhibits NCX1 activity. In the context of Na(+)/K(+)-ATPase (NKA) regulation, pSer-68-PLM is dephosphorylated by protein phosphatase 1 (PP1). PP1 also associates with NCX1; however, the molecular basis of this association is unknown. In this study, we aimed to analyze the mechanisms of PP1 targeting to the NCX1-pSer-68-PLM complex and hypothesized that a direct and functional NCX1-PP1 interaction is a prerequisite for pSer-68-PLM dephosphorylation. Using a variety of molecular techniques, we show that PP1 catalytic subunit (PP1c) co-localized, co-fractionated, and co-immunoprecipitated with NCX1 in rat cardiomyocytes, left ventricle lysates, and HEK293 cells. Bioinformatic analysis, immunoprecipitations, mutagenesis, pulldown experiments, and peptide arrays constrained PP1c anchoring to the K(I/V)FF motif in the first Ca(2+) binding domain (CBD) 1 in NCX1. This binding site is also partially in agreement with the extended PP1-binding motif K(V/I)FF-X5-8Φ1Φ2-X8-9-R. The cytosolic loop of NCX1, containing the K(I/V)FF motif, had no effect on PP1 activity in an in vitro assay. Dephosphorylation of pSer-68-PLM in HEK293 cells was not observed when NCX1 was absent, when the K(I/V)FF motif was mutated, or when the PLM- and PP1c-binding sites were separated (mimicking calpain cleavage of NCX1). Co-expression of PLM and NCX1 inhibited NCX1 current (both modes). Moreover, co-expression of PLM with NCX1(F407P) (mutated K(I/V)FF motif) resulted in the current being completely abolished. In conclusion, NCX1 is a substrate-specifying PP1c regulator protein, indirectly regulating NCX1 activity through pSer-68-PLM dephosphorylation. (© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.)
Databáze:	MEDLINE
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