Pathogenic peptide deviations support a model of adaptive evolution of chordate cardiac performance by troponin mutations

In cardiac muscle, the troponin (cTn) complex is a key regulator of myofilament calcium sensitivity because it serves as a molecular switch required for translating myocyte calcium fluxes into sarcomeric contraction and relaxation. Studies of several species suggest that ectotherm chordates have myofilaments with heightened calcium responsiveness. However, genetic polymorphisms in cTn that cause increased myofilament sensitivity to activating calcium in mammals result in cardiac disease including arrhythmias, diastolic dysfunction, and increased susceptibility to sudden cardiac death. We hypothesized that specific residue modifications in the regulatory arm of troponin I (TnI) were critical in mediating the observed decrease in myofilament calcium sensitivity within the mammalian taxa. We performed large-scale phylogenetic analysis, atomic resolution molecular dynamics simulations and modeling, and computational alanine scanning. This study provides evidence that a His to Ala substitution within mammalian cardiac TnI (cTnI) reduced the thermodynamic potential at the interface between cTnI and cardiac TnC (cTnC) in the calcium-saturated state by disrupting a strong intermolecular electrostatic interaction. This key residue modification reduced myofilament calcium sensitivity by making cTnI molecularly untethered from cTnC. To meet the requirements for refined mammalian adult cardiac performance, we propose that compensatory evolutionary pressures favored mutations that enhanced the relaxation properties of cTn by decreasing its sensitivity to activating calcium.

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Abstract 219: Histidine Substituted Cardiac Troponin I (a164h) Improves Survival And Protects Cardiac Contractility During Acute Hypoxia In The Aged Murine Heart.

Circulation ◽

10.1161/circ.116.suppl_16.ii_23-a ◽

2007 ◽

Vol 116 (suppl_16) ◽

Author(s):

Nathan Palpant ◽

Sharlene Day ◽

Kimber Converso ◽

Joseph Metzger

Keyword(s):

Cardiac Troponin ◽

Troponin I ◽

Acute Hypoxia ◽

Cardiac Contractility ◽

Systolic Pressure ◽

Calcium Sensitivity ◽

The Elderly ◽

Cardiac Troponin I ◽

Pump Failure ◽

Myofilament Calcium Sensitivity

Contractile dysfunction associated with ischemia is a significant cause of morbidity and mortality particularly in the elderly. Strategies designed to protect the aged heart from ischemia-mediated pump failure are needed. We have generated transgenic (Tg) mice expressing a modified form of adult cardiac troponin I, the Ca ++ -activated molecular switch of the myofilament. Consonant with the fetal isoform, this transgene encodes a histidine substitution (A164H) in the critical switch domain of TnI thus increasing myofilament calcium sensitivity in a pH-dependent manner. We hypothesized that aged mice (24 months), intrinsically susceptible to myocardial dysfunction, would retain improved cardiac contractility at baseline and during an acute hypoxic challenge by means of myofilament-mediated calcium sensitization. Methods/Results: At baseline, by echocardiography, Tg hearts had increased systolic function, with a 26% higher mean ejection fraction compared to nontransgenic (Ntg) mice: 75 ± 3% [Tg: n = 13] vs. 63 ± 4% [Ntg: n = 12], P < 0.05, with no differences in diastolic function between the groups. To study the effects of acute hypoxia on cardiac hemodynamics mice underwent microconductance Millar catheterization while ventilated with 12% oxygen. Aged Tg mice had improved survival compared to Ntg mice: time to pump failure (65% of baseline peak systolic pressure) 11.59 ± 1.25 min. [Tg: n = 3] vs. 4.11 ± 1.90 min. [Ntg: n = 3], P < 0.05. After four minutes of hypoxia, Tg mice had markedly improved cardiac contractility compared to Ntg mice with increased stroke volume (30.05 ± 4.49 uL [Tg] vs. 13.23 ± 3.21 uL [Ntg], P < 0.05), end systolic pressure (106.09 ± 11.81 mmHg [Tg] vs. 64.49 ± 4.05 mmHg [Ntg], P < 0.05) and rate of positive left ventricular pressure development (12958.66 ± 2544.68 mmHg/sec [Tg] vs. 5717.00 ± 745.67 mmHg/sec [Ntg], P = 0.05). Conclusion: An alteration in myofilament calcium sensitivity via a pH-responsive histidine button in cardiac troponin I augments baseline heart function in Tg mice over their lifetime. During acute hypoxia, cTnI A164H improves survival in aged mice by maintaining cardiac contractility, and thus holds promise for the design of gene therapeutics to treat pump failure associated with acute ischemic events in the elderly.

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Striated muscle tropomyosin isoforms differentially regulate cardiac performance and myofilament calcium sensitivity

Journal of Muscle Research and Cell Motility ◽

10.1007/s10974-010-9228-3 ◽

2010 ◽

Vol 31 (3) ◽

pp. 227-239 ◽

Cited By ~ 13

Author(s):

Ganapathy Jagatheesan ◽

Sudarsan Rajan ◽

Rafeeq P. H. Ahmed ◽

Natalia Petrashevskaya ◽

Greg Boivin ◽

...

Keyword(s):

Striated Muscle ◽

Calcium Sensitivity ◽

Cardiac Performance ◽

Tropomyosin Isoforms ◽

Muscle Tropomyosin ◽

Myofilament Calcium Sensitivity

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AMP-activated Protein Kinase Phosphorylates Cardiac Troponin I at Ser-150 to Increase Myofilament Calcium Sensitivity and Blunt PKA-dependent Function

Journal of Biological Chemistry ◽

10.1074/jbc.m111.323048 ◽

2012 ◽

Vol 287 (23) ◽

pp. 19136-19147 ◽

Cited By ~ 51

Author(s):

Benjamin R. Nixon ◽

Ariyoporn Thawornkaiwong ◽

Janel Jin ◽

Elizabeth A. Brundage ◽

Sean C. Little ◽

...

Keyword(s):

Protein Kinase ◽

Cardiac Troponin ◽

Troponin I ◽

Calcium Sensitivity ◽

Cardiac Troponin I ◽

Dependent Function ◽

Amp Activated Protein Kinase ◽

Myofilament Calcium Sensitivity

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Frequency-dependent acceleration of relaxation involves decreased myofilament calcium sensitivity

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00778.2006 ◽

2007 ◽

Vol 292 (5) ◽

pp. H2212-H2219 ◽

Cited By ~ 58

Author(s):

Kenneth D. Varian ◽

Paul M. L. Janssen

Keyword(s):

Intracellular Calcium ◽

Calcium Concentration ◽

Troponin I ◽

Zealand White Rabbit ◽

Calcium Sensitivity ◽

Intracellular Calcium Concentration ◽

Calcium Handling ◽

Force Frequency ◽

Frequency Dependent ◽

Myofilament Calcium Sensitivity

The force-frequency relationship is an intrinsic modulator of cardiac contractility and relaxation. Force of contraction increases with frequency, while simultaneously a frequency-dependent acceleration of relaxation occurs. While frequency dependency of calcium handling and sarcoplasmic reticulum calcium load have been well described, it remains unknown whether frequency-dependent changes in myofilament calcium sensitivity occur. We hypothesized that an increase in heart rate that results in acceleration of relaxation is accompanied by a proportional decrease in myofilament calcium sensitivity. To test our hypothesis, ultrathin right ventricular trabeculae were isolated from New Zealand White rabbit hearts and iontophorically loaded with the calcium indicator bis-fura 2. Twitch and intracellular calcium handling parameters were measured and showed a robust increase in twitch force, acceleration of relaxation, and rise in both diastolic and systolic intracellular calcium concentration with increased frequency. Steady-state force-intracellular calcium concentration relationships were measured at frequencies 1, 2, 3, and 4 Hz at 37°C using potassium-induced contractures. EC50 significantly and gradually increased with frequency, from 475 ± 64 nM at 1 Hz to 1,004 ± 142 nM at 4 Hz ( P < 0.05) and correlated with the corresponding changes in half relaxation time. No significant changes in maximal active force development or in the myofilament cooperativity coefficient were found. Myofilament protein phosphorylation was assessed using Pro-Q Diamond staining on protein gels of trabeculae frozen at either 1 or 4 Hz, revealing troponin I and myosin light chain-2 phosphorylation associated with the myofilament desensitization. We conclude that myofilament calcium sensitivity is substantially and significantly decreased at higher frequencies, playing a prominent role in frequency-dependent acceleration of relaxation.

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Myofilament Calcium Sensitivity: Consequences of the Effective Concentration of Troponin I

Frontiers in Physiology ◽

10.3389/fphys.2016.00632 ◽

2016 ◽

Vol 7 ◽

Cited By ~ 13

Author(s):

Jalal K. Siddiqui ◽

Svetlana B. Tikunova ◽

Shane D. Walton ◽

Bin Liu ◽

Meredith Meyer ◽

...

Keyword(s):

Troponin I ◽

Calcium Sensitivity ◽

Effective Concentration ◽

Myofilament Calcium Sensitivity

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Cardiac muscle disease and therapeutic targets

The Journal of General Physiology ◽

10.1085/jgp.2021ecc47 ◽

2021 ◽

Vol 154 (9) ◽

Author(s):

Maike Schuldt ◽

Larissa Dorsch ◽

Diederik Kuster ◽

Jolanda Van der Velden

Keyword(s):

Gene Mutation ◽

Troponin T ◽

Calcium Sensitivity ◽

Cardiac Performance ◽

Muscle Disease ◽

Specific Gene ◽

Tissue Samples ◽

Mutation Carriers ◽

Myofilament Calcium Sensitivity

Hypertrophic cardiomyopathy (HCM) is the most common genetic heart disease. While ∼50% of patients with HCM carry a sarcomere gene mutation (sarcomere mutation-positive, SMP), the genetic background is unknown in the other half of the patients (sarcomere mutation-negative, SMN). Gene mutations are most often present in genes encoding the sarcomere proteins myosin heavy chain, myosin-binding protein C, and troponin T. Studies in cardiac tissue samples from patients with obstructive HCM that were obtained during myectomy surgery showed increased myofilament calcium sensitivity, increased kinetics and tension cost, and a reduction of the super-relaxed state of myosin, which is associated with an energy-conserving status of the crossbridges. The increase in myofilament calcium sensitivity is observed at a low dose of mutant protein, while the magnitude of the increase in calcium sensitivity depends on the specific mutation location. These mutation-mediated myofilament changes may underlie inefficient in vivo cardiac performance in mutation carriers. Reduced cardiac efficiency has been observed before onset of cardiac hypertrophy and at advanced disease stages. In addition, impaired diastolic function is an early disease characteristic of HCM. Our recent proteomics studies revealed increased detyrosination of microtubules, which may be a cause of diastolic dysfunction. Recent treatments that target inefficient cardiac performance, such as myosin inhibitors and metabolic drug therapies, may have the potential to prevent, delay, or even reverse disease in HCM-mutation carriers. Treatment response may depend on the specific gene mutation in SMP individuals and may explain diverse response of HCM patients to therapy. While mutation-mediated cardiomyocyte defects have become clear in past years, more research is warranted to define the cellular pathomechanisms of cardiac dysfunction in SMN patients.

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M8R tropomyosin mutation disrupts actin binding and filament regulation: The beginning affects the middle and end

Journal of Biological Chemistry ◽

10.1074/jbc.ra120.014713 ◽

2020 ◽

Vol 295 (50) ◽

pp. 17128-17137

Author(s):

Alice Ward Racca ◽

Michael J. Rynkiewicz ◽

Nicholas LaFave ◽

Anita Ghosh ◽

William Lehman ◽

...

Keyword(s):

Molecular Dynamics ◽

Thin Filament ◽

Calcium Sensitivity ◽

Actin Binding ◽

Regulatory Function ◽

Azimuthal Position ◽

Dynamics Simulations ◽

And Function ◽

Contraction And Relaxation

Dilated cardiomyopathy (DCM) is associated with mutations in cardiomyocyte sarcomeric proteins, including α-tropomyosin. In conjunction with troponin, tropomyosin shifts to regulate actomyosin interactions. Tropomyosin molecules overlap via tropomyosin–tropomyosin head-to-tail associations, forming a continuous strand along the thin filament. These associations are critical for propagation of tropomyosin's reconfiguration along the thin filament and key for the cooperative switching between heart muscle contraction and relaxation. Here, we tested perturbations in tropomyosin structure, biochemistry, and function caused by the DCM-linked mutation, M8R, which is located at the overlap junction. Localized and nonlocalized structural effects of the mutation were found in tropomyosin that ultimately perturb its thin filament regulatory function. Comparison of mutant and WT α-tropomyosin was carried out using in vitro motility assays, CD, actin co-sedimentation, and molecular dynamics simulations. Regulated thin filament velocity measurements showed that the presence of M8R tropomyosin decreased calcium sensitivity and thin filament cooperativity. The co-sedimentation of actin and tropomyosin showed weakening of actin-mutant tropomyosin binding. The binding of troponin T's N terminus to the actin-mutant tropomyosin complex was also weakened. CD and molecular dynamics indicate that the M8R mutation disrupts the four-helix bundle at the head-to-tail junction, leading to weaker tropomyosin–tropomyosin binding and weaker tropomyosin–actin binding. Molecular dynamics revealed that altered end-to-end bond formation has effects extending toward the central region of the tropomyosin molecule, which alter the azimuthal position of tropomyosin, likely disrupting the mutant thin filament response to calcium. These results demonstrate that mutation-induced alterations in tropomyosin–thin filament interactions underlie the altered regulatory phenotype and ultimately the pathogenesis of DCM.

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