Abstract
Rationale: Myocardial diastolic stiffness and cardiomyocyte passive force (Fpassive) depend in part on titin isoform composition and phosphorylation. Ca2+/calmodulin-dependent protein kinase-IIδ (CaMKIIδ) phosphorylates ion channels, Ca2+-handling and myofilament proteins in the heart, but has not been known to target titin.Objective: To elucidate whether CaMKIIδ phosphorylates titin and regulates Fpassive in normal and failing myocardium.Methods and Results: Titin phosphorylation was assessed in CaMKIIδ/γ double-knockout (DKO) and transgenic CaMKIIδC-overexpressing (TG) mouse hearts, as well as human hearts, by Pro-Q-Diamond/Sypro-Ruby staining, autoradiography, and immunoblotting using phosphospecific titin antibodies. CaMKII-dependent site-specific titin phosphorylation was quantified in vivo by mass spectrometry using SILAC mouse heart tissue mixed with wildtype (WT) or DKO heart. Fpassive of single permeabilized cardiomyocytes was recorded before and after CaMKII administration. We detected hypophosphorylation of all-titin in DKO and hyperphosphorylation in TG compared to WT hearts. Conserved CaMKII-dependent phosphosites were identified within titin's PEVK-domain by quantitative mass spectrometry and confirmed in recombinant human PEVK-fragments. CaMKII also phosphorylated the cardiac titin N2B-unique sequence (N2Bus). Phosphorylation at specific PEVK/N2Bus sites was decreased in DKO and amplified in TG versus WT hearts. Fpassive was elevated in DKO and reduced in TG compared to WT cardiomyocytes. CaMKII-administration lowered Fpassive of WT and DKO cardiomyocytes, an effect blunted by titin-phosphoantibody pretreatment. Human failing hypertrophic hearts revealed higher CaMKII expression/activity and phosphorylation at PEVK/N2Bus sites than nonfailing donor hearts.Conclusions: CaMKIIδ phosphorylates the titin springs at conserved serines/threonines, thereby lowering Fpassive. Deranged CaMKIIδ-dependent titin phosphorylation occurs in heart failure and contributes to altered diastolic stress.
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
