title: Novel role for p90 ribosomal S6 kinase in the regulation of cardiac myofilament phosphorylation.
abstract: In myocardium, the 90-kDa ribosomal S6 kinase (RSK) is activated by diverse stimuli and regulates the sarcolemmal Na(+)/H(+) exchanger through direct phosphorylation. Only limited information is available on other cardiac RSK substrates and functions. We evaluated cardiac myosin-binding protein C (cMyBP-C), a sarcomeric regulatory phosphoprotein, as a potential RSK substrate. In rat ventricular myocytes, RSK activation by endothelin 1 (ET1) increased cMyBP-C phosphorylation at Ser(282), which was inhibited by the selective RSK inhibitor D1870. Neither ET1 nor D1870 affected the phosphorylation status of Ser(273) or Ser(302), cMyBP-C residues additionally targeted by cAMP-dependent protein kinase (PKA). Complementary genetic gain- and loss-of-function experiments, through the adenoviral expression of wild-type or kinase-inactive RSK isoforms, confirmed RSK-mediated phosphorylation of cMyBP-C at Ser(282). Kinase assays utilizing as substrate wild-type or mutated (S273A, S282A, S302A) recombinant cMyBP-C fragments revealed direct and selective Ser(282) phosphorylation by RSK. Immunolabeling with a Ser(P)(282) antibody and confocal fluorescence microscopy showed RSK-mediated phosphorylation of cMyBP-C across the C-zones of sarcomeric A-bands. In chemically permeabilized mouse ventricular muscles, active RSK again induced selective Ser(282) phosphorylation in cMyBP-C, accompanied by significant reduction in Ca(2+) sensitivity of force development and significant acceleration of cross-bridge cycle kinetics, independently of troponin I phosphorylation at Ser(22)/Ser(23). The magnitudes of these RSK-induced changes were comparable with those induced by PKA, which phosphorylated cMyBP-C additionally at Ser(273) and Ser(302). We conclude that Ser(282) in cMyBP-C is a novel cardiac RSK substrate and its selective phosphorylation appears to regulate cardiac myofilament function.
authors.affiliation: King's College London British Heart Foundation Centre, Cardiovascular Division, St Thomas' Hospital, London SE1 7EH, United Kingdom.
authors.forename: Friederike
authors.initials: F
authors.lastname: Cuello
authors.forename: Sonya C
authors.initials: SC
authors.lastname: Bardswell
authors.forename: Robert S
authors.initials: RS
authors.lastname: Haworth
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Ehler
authors.forename: Sakthivel
authors.initials: S
authors.lastname: Sadayappan
authors.forename: Jonathan C
authors.initials: JC
authors.lastname: Kentish
authors.forename: Metin
authors.initials: M
authors.lastname: Avkiran
mesh_terms: D008841:Actin Cytoskeleton / Q000201:enzymology* / Q000235:genetics; D000818:Animals; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D017868:Cyclic AMP-Dependent Protein Kinases / Q000235:genetics / Q000378:metabolism; D006352:Heart Ventricles / Q000166:cytology / Q000201:enzymology*; D051379:Mice; D008822:Mice, Transgenic; D032383:Myocytes, Cardiac / Q000166:cytology / Q000201:enzymology*; D010766:Phosphorylation / Q000502:physiology; D051381:Rats; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000235:genetics / Q000378:metabolism*; D012518:Sarcomeres / Q000201:enzymology*; D000099163:Myosin Binding Protein C
chemical_list: D002352:Carrier Proteins; D017868:Cyclic AMP-Dependent Protein Kinases; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D000099163:Myosin Binding Protein C