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Cardiovasc Res. 2013 Sep 1;99(4):648-56. doi: 10.1093/cvr/cvt144. Epub 2013 Jun 13.

Differential changes in titin domain phosphorylation increase myofilament stiffness in failing human hearts.

Author information

1
Department of Cardiovascular Physiology, Heinrich Heine University Düsseldorf, Germany.

Abstract

AIMS:

Titin-based myofilament stiffness is defined by the expression levels of the cardiac titin-isoforms, N2B and N2BA, and by phosphorylation of the elastic titin domains N2-B unique sequence (N2-Bus) and PEVK. Phosphorylation of the N2-Bus by cGMP-dependent protein kinase (PKG) or cAMP-dependent protein kinase (PKA) decreases titin stiffness, whereas phosphorylation of the PEVK-domain by PKC increases it. We aimed to identify specific sites within the N2-Bus phosphorylated by PKA and PKG and to determine whether differential changes in titin domain phosphorylation could affect passive stiffness in human failing hearts.

METHODS AND RESULTS:

Using mass spectrometry, we identified seven partly conserved PKA/PKG-targeted phosphorylation motifs in human and rat N2-Bus. Polyclonal antibodies to pSer4185, pSer4010, and pSer4099 in the N2-Bus, and to pSer11878 in the PEVK-region were used to quantify titin-domain phosphorylation by western blot analyses of a set of human donor and failing hearts with similar titin-isoform composition. Passive tension determined in skinned human myocardial fibre preparations was significantly increased in failing compared with donor hearts, notably at shorter sarcomere lengths where titin contributes most to total passive tension. Phosphorylation of Ser4185, Ser4010, and Ser4099 in the N2-Bus was significantly reduced in failing hearts, whereas phosphorylation of Ser11878 in the PEVK-region was increased compared with donor hearts.

CONCLUSION:

We conclude that hypo-phosphorylation of the N2-Bus and hyper-phosphorylation of the PEVK domain can act complementary to elevate passive tension in failing human hearts. Differential changes in titin-domain phosphorylation may be important to fine-tune passive myocardial stiffness and diastolic function of the heart.

KEYWORDS:

Connectin; Diastolic dysfunction; Myocardial stiffness; Passive tension; Phospho-specific antibodies

PMID:
23764881
DOI:
10.1093/cvr/cvt144
[Indexed for MEDLINE]
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