Cellular Biology Hyperphosphorylation of Mouse Cardiac Titin Contributes to Transverse Aortic Constriction-Induced Diastolic Dysfunction

Rationale: Mechanisms underlying diastolic dysfunction need to be better understood. Objective: To study the role of titin in diastolic dysfunction using a mouse model of experimental heart failure induced by transverse aortic constriction. titin-based passive stiffness. Changes in titin splicing occur, which lower passive stiffness, but this effect is offset by hyperphosphorylation of residues in titin spring elements, particularly of PEVK S26. Thus, complex changes in titin occur that combined are a major factor in the increased passive myocardial stiffness in HF. A ltered hemodynamics, such as elevated blood pressure and aortic valve diseases, lead to pathological cardiac hypertrophy and ultimately heart failure (HF), characterized by left ventricular dilation, decreased contractility, and increased myocardial stiffness. 1 Although the molecular mechanisms that underlie decreased contractility have been investigated extensively, the molecular basis of diastolic dysfunction is less well-understood. 2– 4 Understanding the mechanisms that govern diastolic dysfunction is important for understanding a wide range of cardiac diseases, including heart failure with preserved ejection fraction (HFpEF), a prevalent disease without effective therapy. 4 –7 The extracel-lular matrix (ECM) is often considered to dominate passive myocardial stiffness, but work focused on the intracellular protein titin has shown that titin is also important for generating passive muscle stiffness. 5 Titin spans from Z-disk to M-line and is extensible in the I-band region of the sarcomere, where it functions as a molecular spring that develops passive force in sarcomeres stretched beyond their slack length (Ϸ1.9 ␮m). 8 Differential splicing results in two main isoforms: the N2B isoform that is stiff (Ϸ3.0 MDa) and the larger N2BA isoform that is more compliant (Ϸ3.4 MDa). 9 These isoforms are coexpressed in the adult heart, with alterations in their expression ratio giving rise to changes in myocardial stiffness. 10 In addition, titin phosphorylation also alters myocardial stiffness. Both protein kinase A (PKA) 11 and protein kinase G (PKG) 12 phosphorylate titin (most likely at the same site in the N2B spring element 12) and reduce myocardial stiffness (increased compliance). Evidence suggests that hypophosphorylation of PKA/PKG sites on titin contribute to reduced compliance in HFpEF patients. 13 Recently, it was found that titin is also a target of protein kinase C (PKC)␣, and that PKC␣ phosphor-ylation of two conserved serines in the proline glutamate valine lysine (PEVK) region of titin (S26 and S170 in the PEVK of the N2B cardiac titin isoform) increases cardiac stiffness. 14 –16 Here, we …