CIC   05421
CENTRO DE INVESTIGACIONES CARDIOVASCULARES "DR. HORACIO EUGENIO CINGOLANI"
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
The multifunctional calcium/calmodulin-dependent protein kinase II delta (CaMKIId) phosphorylates titin spring element
Autor/es:
226. HIDALGOC, METHAWASIN M, CHUNG CH S1, BOGOMOLOVAS J, GASCH A, LABEIT S, MATTIAZZI A. GRANZIER H
Lugar:
San Diego
Reunión:
Congreso; 56th meeting of the Biophysical Society; 2012
Institución organizadora:
Biophysical Society
Resumen:
Titin-based passive stiffness is post-translationally regulated by several kinases that phosphorylate specific spring elements located within titinLs elastic I-band region. Whether titin is phosphorylated by calcium/calmodulin dependent protein kinase II (CaMKII), an important regulator of cardiac function and disease, has not been addressed. The aim of this work was to determine whether CaMKIIƒÂ, the predominant CaMKII isoform in the heart, phosphorylates titin, and to use phosphorylation assays and mass spectrometry to study which of titinLs spring elements might be targeted by CaMKIIƒÂ. It was found that CaMKIIƒÂ phosphorylates titin in mouse LV skinned fibers, that the CaMKIIƒÂ sites can be dephosphorylated by protein phosphatase 1 (PP1), and that under baseline conditions, in both intact isolated hearts and skinned myocardium, about half of the CaMKIIƒÂ sites are phosphorylated. Mass spectrometry revealed that both the N2B and PEVK segments are targeted by CaMKIIƒÂ at several conserved serine residues. Whether phosphorylation of titin by CaMKIIƒÂ occurs in vivo, was tested in several conditions using back phosphorylation assays and phospho-specific antibodies to CaMKIIƒÂ sites. Reperfusion following global ischemia increased the phosphorylation level of CaMKIIƒÂ sites on titin and this effect was abolished by the CaMKII inhibitor KN-93. No changes in the phosphorylation level of the PEVK element were found suggesting that the increased phosphorylation level of titin in IR (ischemia reperfusion) might be due to phosphorylation of the N2B element. The findings of these studies show for the first time that titin can be phosphoryalated by CaMKIIƒÂ, both in vitro and in vivo, and that titinLs molecular spring region that determines diastolic stiffness is a target of CaMKIIƒÂ.
rds']