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Henk L Granzier

Publications and source records attributed to Henk L Granzier.

12 recordsLinked to original sources
PubMed2006

The giant muscle protein titin is an adjustable molecular spring.

When muscles are stretched, the giant protein titin develops passive force. Titin's force performs important functions that include maintaining the structural integrity of the sarcomere, and triggering signal transduction pathways. We propose that the mechanical properties of titin can be tuned according to the mechanical demands places on muscle, using mechanisms that include alternative splicing and posttranslational modifications.

Calcium↗
PubMed2005

Titin and diaphragm dysfunction in chronic obstructive pulmonary disease.

RATIONALE: Recently, we have shown that Ca2+-activated force generation in diaphragm single fibers is impaired in patients with mild to moderate chronic obstructive pulmonary disease (COPD). For optimal active-force generation, the passive elasticity provided by titin is indispensable. OBJECTIVES: In the present study, we determined the passive-tension-length relations of single fibers of patients with mild to moderate COPD, hypothesizing that passive-elastic properties of diaphragm fibers are compromised. METHODS: Passive-tension-length relations were determined in diaphragm fibers from patients with and without COPD (predicted mean FEV1, 76 and 102%, respectively). In diaphragm homogenates titin expression was studied at the protein level by gel electrophoresis and at the transcript level by using a novel titin exon microarray. RESULTS: Diaphragm fibers from patients with COPD generate less passive tension on stretch. Titin content in the diaphragm did not differ between patients with and without COPD. However, titin exon transcript studies revealed up-regulation of seven exons, which code for spring elements in the elastic segment rich in proline, glutamate, valine, and lysine. Immunofluorescence analysis indicated elevated protein expression of the up-regulated splice variant in the COPD diaphragm. Simulation studies on titin molecules including the amino acids encoded by the seven up-regulated exons predicted reduced passive-tension generation on molecule stretch. CONCLUSIONS: Passive-tension generation of diaphragm single fibers is reduced in patients with COPD. Our results suggest that alternative splicing of the titin gene, resulting in increased length of the elastic segment rich in proline, glutamate, valine, and lysine, is involved. Interestingly, these changes occur already in patients with mild to moderate COPD.

Adult↗
PubMed2005

Titin/connectin-based modulation of the Frank-Starling mechanism of the heart.

The basis of the Frank-Starling mechanism of the heart is the increase in active force when muscle is stretched. Various findings have shown that muscle length, i.e., sarcomere length (SL), modulates activation of cardiac myofilaments at a given concentration of Ca2+ ([Ca2+]). This augmented Ca2+ activation with SL, commonly known as "length-dependent activation", is manifested as the leftward shift of the force-pCa (= -log [Ca2+]) relation as well as by the increase in maximal Ca2+ -activated force. Despite the numerous studies that have been undertaken, the molecular mechanism(s) of length-dependent activation is (are) still not fully understood. The giant sarcomere protein titin/connectin is the largest protein known to date. Titin/connectin is responsible for most passive force in vertebrate striated muscle and also functions as a molecular scaffold during myofibrillogenesis. Recent studies suggest that titin/connectin plays an important role in length-dependent activation by sensing stretch and promoting actomyosin interaction. Here we review and extend this previous work and focus on the mechanism by which titin/connectin might modulate actomyosin interaction.

Actomyosin↗
PubMed2005

Phosphorylation of titin modulates passive stiffness of cardiac muscle in a titin isoform-dependent manner.

We investigated the effect of protein kinase A (PKA) on passive force in skinned cardiac tissues that express different isoforms of titin, i.e., stiff (N2B) and more compliant (N2BA) titins, at different levels. We used rat ventricular (RV), bovine left ventricular (BLV), and bovine left atrial (BLA) muscles (passive force: RV > BLV > BLA, with the ratio of N2B to N2BA titin, approximately 90:10, approximately 40:60, and approximately 10:90%, respectively) and found that N2B and N2BA isoforms can both be phosphorylated by PKA. Under the relaxed condition, sarcomere length was increased and then held constant for 30 min and the peak passive force, stress-relaxation, and steady-state passive force were determined. Following PKA treatment, passive force was significantly decreased in all muscle types with the effect greatest in RV, lowest in BLA, and intermediate in BLV. Fitting the stress-relaxation data to the sum of three exponential decay functions revealed that PKA blunts the magnitude of stress-relaxation and accelerates its time constants. To investigate whether or not PKA-induced decreases in passive force result from possible alteration of titin-thin filament interaction (e.g., via troponin I phosphorylation), we conducted the same experiments using RV preparations that had been treated with gelsolin to extract thin filaments. PKA decreased passive force in gelsolin-treated RV preparations with a magnitude similar to that observed in control preparations. PKA was also found to decrease restoring force in skinned ventricular myocytes of the rat that had been shortened to below the slack length. Finally, we investigated the effect of the beta-adrenergic receptor agonist isoprenaline on diastolic force in intact rat ventricular trabeculae. We found that isoprenaline phosphorylated titin and that it reduced diastolic force to a degree similar to that found in skinned RV preparations. Taken together, these results suggest that during beta-adrenergic stimulation, PKA increases ventricular compliance in a titin isoform-dependent manner.

Adrenergic beta-Agonists↗
PubMed2004

Altered titin expression, myocardial stiffness, and left ventricular function in patients with dilated cardiomyopathy.

BACKGROUND: The role of the giant protein titin in patients with heart failure is not well established. We investigated titin expression in patients with end-stage heart failure resulting from nonischemic dilated cardiomyopathy, in particular as it relates to left ventricular (LV) myocardial stiffness and LV function. METHODS AND RESULTS: SDS-agarose gels revealed small N2B (stiff) and large N2BA (compliant) cardiac titin isoforms with a mean N2BA:N2B expression ratio that was significantly (P<0.003) increased in 20 heart failure patients versus 6 controls. However, total titin was unchanged. The coexpression ratio was highest in a subsample of patients with an impaired LV relaxation pattern (n=7), intermediate in those with pseudonormal filling (n=6), and lowest in the group with restrictive filling (n=7). Mechanical measurements on LV muscle strips dissected from these hearts (n=8) revealed that passive muscle stiffness was significantly reduced in patients with a high N2BA:N2B expression ratio. Clinical correlations support the relevance of these changes for LV function (assessed by invasive hemodynamics and Doppler echocardiography). A positive correlation between the N2BA:N2B titin isoform ratio and deceleration time of mitral E velocity, A wave transit time, and end diastolic volume/pressure ratio was found. These changes affect exercise tolerance, as indicated by the positive correlation between the N2BA:N2B isoform ratio and peak O2 consumption (n=10). Upregulated N2BA expression was accompanied by increased expression levels of titin-binding proteins (cardiac ankyrin repeat protein, ankrd2, and diabetes ankyrin repeat protein) that bind to the N2A element of N2BA titin (studied in 13 patients). CONCLUSIONS: Total titin content was unchanged in end-stage failing hearts and the more compliant N2BA isoform comprised a greater percentage of titin in these hearts. Changes in titin isoform expression in heart failure patients with dilated cardiomyopathy significantly impact diastolic filling by lowering myocardial stiffness. Upregulation of titin-binding proteins indicates that the importance of altered titin expression might extend to cell signaling and regulation of gene expression.

Adult↗
PubMed2004

The giant protein titin: a major player in myocardial mechanics, signaling, and disease.

The sarcomere contains, in addition to thin and thick filaments, a filament composed of the giant protein titin (also known as connectin). Titin molecules anchor in the Z-disc and extend to the M-line region of the sarcomere. The majority of titin's I-band region functions as a molecular spring. This spring maintains the precise structural arrangement of thick and thin filaments, and gives rise to passive muscle stiffness; an important determinant of diastolic filling. Earlier work on titin has been reviewed before. In this study, our main focus is on recent findings vis-à-vis titin's molecular spring segments in cardiac titins, including the discovery of fetal cardiac isoforms with novel spring elements. We also discuss new insights regarding the role of titin as a biomechanical sensor and signaling molecule. We will end with focusing on the rapidly growing knowledge regarding titinopathies.

Animals↗
PubMed2004

Cellular and molecular mechanisms of systolic and diastolic dysfunction in an avian model of dilated cardiomyopathy.

We investigated the cellular and molecular mechanisms of systolic and diastolic dysfunction in a furazolidone (Fz)-induced model of dilated cardiomyopathy (DCM) in turkey poults. Serial echocardiograms disclosed marked systolic dysfunction in the Fz-treated poults, and ventricular weight and left ventricular (LV)/body weight ratio were significantly increased. Isolated heart experiments were performed to determine LV pressure-volume (P-V) relationships. In addition, LV sarcomere lengths (SLs) were measured after hearts had been fixed, and wall stress (sigma)-SL relationships were determined. When compared to control hearts, LV chamber volume in DCM hearts was approximately 3-fold increased, the active or developed LV P-V relationship was markedly depressed, the passive or diastolic P-V relationship was steeper, and SLs were significantly shorter. However, the developed sigma-SL relationships of DCM and control hearts were not different indicating that intrinsic myocardial capacity to generate active force is unaffected in this model of DCM. In contrast, passive sigma, and passive tension in trabecular muscle preparations increased much more steeply with SL in DCM than normal hearts. Trabecular muscle experiments disclosed that the increase in passive myocardial stiffness was primarily collagen based. Titin, the giant sarcomeric molecule, which is an important determinant of passive myocyte properties in normal myocardium, did not contribute significantly to increased passive myocardial stiffness in DCM. We conclude that increased collagen-based passive myocardial stiffness is the major cause of the steeper passive or diastolic P-V relationship in DCM. Further, altered passive myocardial properties and ventricular geometry in DCM play a critical role to reduce ventricular systolic function by limiting SL extension during diastole, thereby limiting the use of the myocardial length-tension relationship.

Animals↗
PubMed2004

Titin isoform-dependent effect of calcium on passive myocardial tension.

We studied the effects of Ca2+ on titin (connectin)-based passive tension in skinned myocardium expressing either predominantly N2B titin (rat right ventricle, RRV) or predominantly N2BA titin (bovine left atrium, BLA). Actomyosin-based tension was abolished to undetectably low levels by selectively removing the thin filaments with a Ca2+-insensitive gelsolin fragment (FX-45). Myocardium was stretched in the presence and absence of Ca2+, and passive tension was measured. Ca2+ significantly increased passive tension during and after stretch in the BLA. The increase was insensitive to the actomyosin inhibitor 2,3-butanedione 2-monoxime, supporting the conclusion that the effect is titin based. Passive tension did not respond to calcium in the RRV, indicating that passive tension developed by N2B titin is calcium insensitive. Western blot analysis and immunofluorescence studies indicated that N2BA titin expresses E-rich PEVK motifs, whereas they are absent from N2B titin, supporting earlier single molecule studies that reported that E-rich motifs are required for calcium sensitivity. We conclude that calcium affects passive myocardial tension in a titin isoform-dependent manner.

Actin Cytoskeleton↗
PubMed2003

Mechanics and structure of titin oligomers explored with atomic force microscopy.

Titin is a giant polypeptide that spans half of the striated muscle sarcomere and generates passive force upon stretch. To explore the elastic response and structure of single molecules and oligomers of titin, we carried out molecular force spectroscopy and atomic force microscopy (AFM) on purified full-length skeletal-muscle titin. From the force data, apparent persistence lengths as long as approximately 1.5 nm were obtained for the single, unfolded titin molecule. Furthermore, data suggest that titin molecules may globally associate into oligomers which mechanically behave as independent wormlike chains (WLCs). Consistent with this, AFM of surface-adsorbed titin molecules revealed the presence of oligomers. Although oligomers may form globally via head-to-head association of titin, the constituent molecules otherwise appear independent from each other along their contour. Based on the global association but local independence of titin molecules, we discuss a mechanical model of the sarcomere in which titin molecules with different contour lengths, corresponding to different isoforms, are held in a lattice. The net force response of aligned titin molecules is determined by the persistence length of the tandemly arranged, different WLC components of the individual molecules, the ratio of their overall contour lengths, and by domain unfolding events. Biased domain unfolding in mechanically selected constituent molecules may serve as a compensatory mechanism for contour- and persistence-length differences. Variation in the ratio and contour length of the component chains may provide mechanisms for the fine-tuning of the sarcomeric passive force response.

Adsorption↗