title: Role of dual-site phospholamban phosphorylation in the stunned heart: insights from phospholamban site-specific mutants.
abstract: Phosphorylation of phospholamban (PLB) at Ser16 (protein kinase A site) and at Thr17 [Ca2+/calmodulin kinase II (CaMKII) site] increases sarcoplasmic reticulum Ca2+ uptake and myocardial contractility and relaxation. In perfused rat hearts submitted to ischemia-reperfusion, we previously showed an ischemia-induced Ser16 phosphorylation that was dependent on beta-adrenergic stimulation and an ischemia and reperfusion-induced Thr17 phosphorylation that was dependent on Ca2+ influx. To elucidate the relationship between these two PLB phosphorylation sites and postischemic mechanical recovery, rat hearts were submitted to ischemia-reperfusion in the absence and presence of the CaMKII inhibitor KN-93 (1 microM) or the beta-adrenergic blocker dl-propranolol (1 microM). KN-93 diminished the reperfusion-induced Thr17 phosphorylation and depressed the recovery of contraction and relaxation after ischemia. dl-Propranolol decreased the ischemia-induced Ser16 phosphorylation but failed to modify the contractile recovery. To obtain further insights into the functional role of the two PLB phosphorylation sites in postischemic mechanical recovery, transgenic mice expressing wild-type PLB (PLB-WT) or PLB mutants in which either Thr17 or Ser16 were replaced by Ala (PLB-T17A and PLB-S16A, respectively) into the PLB-null background were used. Both PLB mutants showed a lower contractile recovery than PLB-WT. However, this recovery was significantly impaired all along reperfusion in PLB-T17A, whereas it was depressed only at the beginning of reperfusion in PLB-S16A. Moreover, the recovery of relaxation was delayed in PLB-T17A, whereas it did not change in PLB-S16A, compared with PLB-WT. These findings indicate that, although both PLB phosphorylation sites are involved in the mechanical recovery after ischemia, Thr17 appears to play a major role.
authors.affiliation: Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Médicas 60 y 120, 1900 La Plata, Argentina. ramattia@atlas.med.unlp.edu.ar
authors.forename: M
authors.initials: M
authors.lastname: Said
authors.forename: L
authors.initials: L
authors.lastname: Vittone
authors.forename: C
authors.initials: C
authors.lastname: Mundina-Weilenmann
authors.forename: P
authors.initials: P
authors.lastname: Ferrero
authors.forename: E G
authors.initials: EG
authors.lastname: Kranias
authors.forename: A
authors.initials: A
authors.lastname: Mattiazzi
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D002135:Calcium-Binding Proteins / Q000235:genetics* / Q000378:metabolism*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D009200:Myocardial Contraction / Q000502:physiology; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000503:physiopathology; D017682:Myocardial Stunning / Q000378:metabolism* / Q000503:physiopathology*; D010766:Phosphorylation; D051381:Rats; D017208:Rats, Wistar; D000099211:Phospholamban
chemical_list: D002135:Calcium-Binding Proteins; D000099211:Phospholamban