Targeted deletion of titin N2B region leads to diastolic dysfunction and cardiac atrophy

Abstract

Titin is a giant protein that is in charge of the assembly and passive mechanical properties of the sarcomere. Cardiac titin contains a unique N2B region, which has been proposed to modulate elasticity of the titin filament and to be important for hypertrophy signaling and the ischemic stress response through its binding proteins FHL2 and ␣B-crystallin, respectively. To study the role of the titin N2B region in systole and diastole of the heart, we generated a knockout (KO) mouse deleting only the N2B exon 49 and leaving the remainder of the titin gene intact. The resulting mice survived to adulthood and were fertile. Although KO hearts were small, they produced normal ejection volumes because of an increased ejection fraction. FHL2 protein levels were significantly reduced in the KO mice, a finding consistent with the reduced size of KO hearts. Ultrastructural analysis revealed an increased extension of the remaining spring elements of titin (tandem Ig segments and the PEVK region), which, together with the reduced sarcomere length and increased passive tension derived from skinned cardiomyocyte experiments, translates to diastolic dysfunction as documented by echocardiography. We conclude from our work that the titin N2B region is dispensable for cardiac development and systolic properties but is important to integrate trophic and elastic functions of the heart. The N2B-KO mouse is the first titin-based model of diastolic dysfunction and, considering the high prevalence of diastolic heart failure, it could provide future mechanistic insights into the disease process.cardiac muscle ͉ hypertrophy ͉ mechanics ͉ cardiology ͉ disease T itin forms a continuous filament along the myofibril that determines the elastic properties of cardiac myocytes (for review, see ref. 1). The extensible region of titin is found in the I-band region of the sarcomere and comprises tandemly arranged Ig-like domains and the so-called PEVK region (2). In addition, cardiac titin contains a third extensible region, the N2B element (2), which is absent in skeletal muscle. The N2B region extends greatly toward the upper limit of the physiological sarcomere length of cardiac muscle (3, 4). It has been suggested that this extension reduces the steepness of the passive forcesarcomere length relation, decreasing the likelihood of the unfolding of Ig domains (3). Mutations in the N2B region can lead to dilated or hypertrophic cardiomyopathy, apparently through altered affinity to FHL2, a heart-specific member of the LIM domain gene family (5). To understand the role of the titin N2B region in cardiac function and disease, we have eliminated exon 49, which encodes the N2B region, and investigated its effect on the mechanical and trophical properties of the knockout (KO) heart.

Dystrophin-deficient cardiomyopathy in mouse: Expression of Nox4 and Lox are associated with fibrosis and altered functional parameters in the heart

“…[26] Our results using high frequency echocardiography for the measurement of shortening fraction are also consistent with other studies using similar techniques in different mouse models. [27], [28], [29], [30] The calculation of ejection fraction is also consistent with a recent study. [31] The additional capability of directly measuring percent ejection fraction in the 2D parasternal long axis allows high frequency echocardiography to be used alongside other newer imaging modalities such as 3D echo and MRI.…”

Deleting titin’s I-band/A-band junction reveals critical roles for titin in biomechanical sensing and cardiac function

“…The PEVK KO and IG KO models (39,40) both share with the Ttn ΔIAjxn model (i) increased N2B strain, (ii) increased FHL2 expression, and (iii) LV hypertrophy. Additionally, FHL2 expression is greatly reduced in the N2B KO mouse in which the N2B element has been deleted and hearts are atrophied (41). Hence, we propose that both global chronic isoproterenol activation (38) and local increased strain of titin's N2B element (e.g.…”

Titin‐based mechanosensing modulates muscle hypertrophy

“…Titin‐based mechanosensing might involve titin‐binding proteins that form several signalling ‘hubs’ along the titin molecule. In line with a mechanosensing role for titin, disruption of two of these hubs depresses muscle gene expression 3, 4. Furthermore, decreasing titin stiffness in the heart blunts hypertrophy development during mechanical overload6 and increasing titin's stiffness has the opposite effect,7, 8 processes that presumably involve titin‐binding proteins (e.g.…”