title: CaMKII in addition to MLCK contributes to phosphorylation of regulatory light chain in cardiomyocytes.
abstract: The aim was to identify kinase activities involved in the phosphorylation of regulatory light chain (RLC) in situ in cardiomyocytes. In electrically stimulated rat cardiomyocytes, phosphatase inhibition by calyculin A unmasked kinase activities evoking an increase of phosphorylated RLC (P-RLC) from about 16% to about 80% after 80 min. The phosphorylation rate in cardiomyocytes was reduced by about 40% by the myosin light chain kinase (MLCK) inhibitor, ML-7. In rat ventricular muscle strips, calyculin A induced a positive inotropic effect that correlated with P-RLC levels. The inotropic effect and P-RLC elevation were abolished by ML-7 treatment. The kinase activities phosphorylating RLC in cardiomyocytes were reduced by about 60% by the non-selective kinase inhibitor staurosporine and by about 50% by the calmodulin antagonist W7. W7 eliminated the inhibitory effect of ML-7, suggesting that the cardiac MLCK is Ca(2+)/calmodulin (CaM)-dependent. The CaM-dependent kinase II (CaMKII) inhibitor KN-93 attenuated the calyculin A-induced RLC phosphorylation by about 40%, indicating a contribution from CaMKII. The residual phosphorylation in the presence of W7 indicated that also CaM-independent kinase activities might contribute. RLC phosphorylation was insensitive to protein kinase C inhibition. In conclusion, in addition to MLCK, CaMKII phosphorylates RLC in cardiomyocytes. Involvement of other kinases cannot be excluded.
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: hilde.eikemo@ntnu.no.
authors.forename: Hilde
authors.initials: H
authors.lastname: Eikemo
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: l.r.moltzau@medisin.uio.no.
authors.forename: Lise Román
authors.initials: LR
authors.lastname: Moltzau
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: rizwan.hussain@medisin.uio.no.
authors.forename: Rizwan I
authors.initials: RI
authors.lastname: Hussain
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway.
authors.forename: Cam H T
authors.initials: CH
authors.lastname: Nguyen
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: eirik.qvigstad@medisin.uio.no.
authors.forename: Eirik
authors.initials: E
authors.lastname: Qvigstad
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: f.o.levy@medisin.uio.no.
authors.forename: Finn Olav
authors.initials: FO
authors.lastname: Levy
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: tor.skomedal@medisin.uio.no.
authors.forename: Tor
authors.initials: T
authors.lastname: Skomedal
authors.affiliation: Department of Pharmacology, Institute of Clinical Medicine, University of Oslo and Oslo University Hospital, Oslo, Norway; K.G. Jebsen Cardiac Research Centre and Center for Heart Failure Research, Faculty of Medicine, University of Oslo, Oslo, Norway. Electronic address: j.b.osnes@medisin.uio.no.
authors.forename: Jan-Bjørn
authors.initials: JB
authors.lastname: Osnes
mesh_terms: D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2 / Q000378:metabolism*; D002478:Cells, Cultured; D004789:Enzyme Activation; D005786:Gene Expression Regulation / Q000502:physiology; D008297:Male; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000502:physiology*; D018994:Myosin Light Chains / Q000378:metabolism*; D009219:Myosin-Light-Chain Kinase / Q000378:metabolism*; D010766:Phosphorylation / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar
chemical_list: D018994:Myosin Light Chains; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D009219:Myosin-Light-Chain Kinase
keywords: Ca(2+)/calmodulin-dependent kinase II (CaMKII); MLC2); Myosin light chain 2; Myosin light chain kinase (MLCK); Phosphatase inhibition; Phosphorylation; Regulatory light chain (RLC