Pathogenesis associated with a restrictive cardiomyopathy mutant in cardiac troponin T is due to reduced protein stability and greatly increased myofilament Ca2+ sensitivity

Parvatiyar, Michelle S.; Pinto, José R.

doi:10.1016/j.bbagen.2014.09.029

Cited by 7 publications

(6 citation statements)

References 61 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For example, mutations in the TnT gene are one of the predominant causes of hypertrophy ( Seidman and Seidman 2001 ; Di Pasquale et al 2012 ). Most of these TnT mutations exhibit increased calcium sensitivity and activation of muscle contractility ( Harada and Potter 2004 ; Parvatiyar and Pinto 2015 ; Gilda et al 2016 ), a similar pathogenesis to the hypercontraction produced by TnT mutants in Drosophila ( Nongthomba et al 2003 , 2007 ). Viswanathan et al (2014 ) have shown that the up 101 mutation generates a muscle abnormality similar to human cardiomyopathy through sensitive calcium regulation in the Drosophila heart.…”

Section: Discussionmentioning

confidence: 96%

Overexpression of miRNA-9 Generates Muscle Hypercontraction Through Translational Repression of Troponin-T inDrosophila melanogasterIndirect Flight Muscles

Katti

Thimmaya

Madan

et al. 2017

G3 Genes|Genomes|Genetics

View full text Add to dashboard Cite

MicroRNAs (miRNAs) are small noncoding endogenous RNAs, typically 21–23 nucleotides long, that regulate gene expression, usually post-transcriptionally, by binding to the 3′-UTR of target mRNA, thus blocking translation. The expression of several miRNAs is significantly altered during cardiac hypertrophy, myocardial ischemia, fibrosis, heart failure, and other cardiac myopathies. Recent studies have implicated miRNA-9 (miR-9) in myocardial hypertrophy. However, a detailed mechanism remains obscure. In this study, we have addressed the roles of miR-9 in muscle development and function using a genetically tractable model system, the indirect flight muscles (IFMs) of Drosophila melanogaster. Bioinformatics analysis identified 135 potential miR-9a targets, of which 27 genes were associated with Drosophila muscle development. Troponin-T (TnT) was identified as major structural gene target of miR-9a. We show that flies overexpressing miR-9a in the IFMs have abnormal wing position and are flightless. These flies also exhibit a loss of muscle integrity and sarcomeric organization causing an abnormal muscle condition known as “hypercontraction.” Additionally, miR-9a overexpression resulted in the reduction of TnT protein levels while transcript levels were unaffected. Furthermore, muscle abnormalities associated with miR-9a overexpression were completely rescued by overexpression of TnT transgenes which lacked the miR-9a binding site. These findings indicate that miR-9a interacts with the 3′-UTR of the TnT mRNA and downregulates the TnT protein levels by translational repression. The reduction in TnT levels leads to a cooperative downregulation of other thin filament structural proteins. Our findings have implications for understanding the cellular pathophysiology of cardiomyopathies associated with miR-9 overexpression.

show abstract

Section: Discussionmentioning

confidence: 96%

Overexpression of miRNA-9 Generates Muscle Hypercontraction Through Translational Repression of Troponin-T inDrosophila melanogasterIndirect Flight Muscles

Katti

Thimmaya

Madan

et al. 2017

G3 Genes|Genomes|Genetics

View full text Add to dashboard Cite

show abstract

“…As the antiphosphorylated protein antibodies were relatively specific for the phosphorylated molecules though a low levels of signal of λ phosphatase-treated proteins was detected by these antibodies (Online Figure V), these results suggest that under basal condition in the normal mouse heart, relatively high PKA activity at the myofilament, and longitudinal SR compartments caused relatively high levels of phosphorylation for PLB, cMyBP-C, and TnI. Another possibility was that there could be more phosphatase activity in these cellular compartments because the loss of basal PKA activity reduced the phosphorylation of some protein phosphatase inhibitors (eg, ARPP19 (cAMP-regulated phosphoprotein 19) and α-endosulfine [Online Table I]) and thus their ability to inhibit phosphatases.…”

Section: Pka Activity Suppression Differentially Alter Basal Level Ph...mentioning

confidence: 95%

“…18 The cTnT-ΔE96 mutant causes restrictive cardiomyopathy probably by enhancing myofilament Ca 2+ sensitivity that cannot be reduced by PKA phosphorylation. 19 What is not clear is if these disease-related changes in PKA signaling are causes or effects of heart disease.…”

Section: Novelty and Significancementioning

confidence: 99%

“…1 In chronic cardiovascular diseases that increase the work demands of the heart, there is chronic activation of this signaling pathway with reduced PKA-mediated responses. 41 Many studies have also shown that reduced or loss of PKAdependent phosphorylation of myofilament protein, 16,18,19 Cav1.2 α1c 42 and PLB 17 due to genetic mutations/manipulations of the PKA phosphorylation sites and reduced PKA activity leads to cardiomyopathy. However, the contribution of depressed PKA activity in disease has not been clearly defined, in large part because of the lack of specific and cardiac-targeted PKA inhibitors 20 or cardiac-specific PKA knockout mice.…”

Section: The Loss Of Pka Activity and Cardiac Dysfunctionmentioning

confidence: 99%

See 1 more Smart Citation

Cardiomyocyte PKA Ablation Enhances Basal Contractility While Eliminates Cardiac β-Adrenergic Response Without Adverse Effects on the Heart

Zhang

Wang

Zhang

et al. 2019

Circulation Research

View full text Add to dashboard Cite

Rationale: PKA (Protein Kinase A) is a major mediator of β-AR (β-adrenergic) regulation of cardiac function, but other mediators have also been suggested. Reduced PKA basal activity and activation are linked to cardiac diseases. However, how complete loss of PKA activity impacts on cardiac physiology and if it causes cardiac dysfunction have never been determined. Objectives: We set to determine how the heart adapts to the loss of cardiomyocyte PKA activity and if it elicits cardiac abnormalities. Methods and Results: (1) Cardiac PKA activity was almost completely inhibited by expressing a PKA inhibitor peptide in cardiomyocytes (cPKAi) in mice; (2) cPKAi reduced basal phosphorylation of 2 myofilament proteins (TnI [troponin I] and cardiac myosin binding protein C), and one longitudinal SR (sarcoplasmic reticulum) protein (PLB [phospholamban]) but not of the sarcolemmal proteins (Cav1.2 α1c and PLM [phospholemman]), dyadic protein RyR2, and nuclear protein CREB (cAMP response element binding protein) at their PKA phosphorylation sites; (3) cPKAi increased the expression of CaMKII (Ca 2+ /calmodulin-dependent kinase II), the Cav1.2 β subunits and current, but decreased CaMKII phosphorylation and CaMKII-mediated phosphorylation of PLB and RyR2; (4) These changes resulted in significantly enhanced myofilament Ca 2+ sensitivity, prolonged contraction, slowed relaxation but increased myocyte Ca 2+ transient and contraction amplitudes; (5) Isoproterenol-induced PKA and CaMKII activation and their phosphorylation of proteins were prevented by cPKAi; (6) cPKAi abolished the increases of heart rate, and cardiac and myocyte contractility by a β-AR agonist (isoproterenol), showing an important role of PKA and a minimal role of PKA-independent β-AR signaling in acute cardiac regulation; (7) cPKAi mice have partial exercise capability probably by enhancing vascular constriction and ventricular filling during β-AR stimulation; and (8) cPKAi mice did not show any cardiac functional or structural abnormalities during the 1-year study period. Conclusions: PKA activity suppression induces a unique Ca 2+ handling phenotype, eliminates β-AR regulation of heart rates and cardiac contractility but does not cause cardiac abnormalities.

show abstract

“…TnT is thought to contribute to cooperative activation of force development via its interaction and influence on Tm (46). cTnT is among the most frequently mutated genes in DCM (26, 45), and mutations of cTnT have also been associated with the development of HCM (75,141) and restrictive cardiomyopathy (RCM) (36,119).…”

Section: 2b Troponin Complexmentioning

confidence: 99%

Molecular regulation of sarcomere function

Robinett¹

View full text Add to dashboard Cite

Molecular regulation of striated muscle contraction is widely thought to be regulated through canonical thin filament mechanisms, i.e., Ca2+ activation of thin filaments. Dual filament regulation is a novel model of striated muscle contraction, in which thick filament activation is regulated by the stress, i.e., thick filaments are activated when the muscle bears a relatively high external load. This study aimed to provide insight on the molecular regulation of multiple biophysical properties of striated muscle. Myosin binding protein-C (MyBP-C) is capable of binding both thin and thick filaments rendering it a well suited regulatory molecule. These interactions are modulated by MyBP-C phosphorylation. MyBP-C's interaction with myosin has the potential to sequester myosin heads in the OFF state and genetic mutations of three myosin binding residues in cMyBP-C causes cardiac hypertrophy. We investigated MyBP-C's regulation of myofilament/sarcomere function by performing slack-restretch protocol to test force development and transient force overshoot and step-stretch protocol to test stretch activation using PKA and lambda phosphatase to modulate sMyBP-C phosphorylation. PKA doubled the transient force overshoot at low (~25 percent) Ca2+ activations in slow-twitch skeletal muscle fibers, while lambda phosphatase decreased transient force overshoot at low (~25 percent) and medium (~50 percent) Ca2+ activation levels. Rates of force development were increased following PKA and decreased following lambda phosphatase at all Ca2+ activation levels. Similarly, PKA increased stretch activation at low (~25 percent) Ca2+ activation levels, while rates of delayed force development (kdf) increased at all Ca2+ activation levels. Additionally, MyBP-C is localized in striated muscle throughout the inner third of each half thick filament, a region known as the C-zone. MyBP-C is capable of binding actin with micromolar affinity. In isolated filaments, thin filament sliding slows as they transverse the C-zone of thick filaments. In support of this, extraction or genetic ablation of MyBP-C sped rates of sarcomere shortening, i.e., MyBP-C has the potential to impose an internal drag that impedes loaded shortening. However, this has not been shown at the sarcomere level; therefore, we designed experiments whereby permeabilized slow-twitch skeletal muscle fibers were stretched to SLs at which the thin filaments were outside of the C-zone (SL ~3.08-3.04 [mu letter]m), then sub-isometric force clamps were performed to elicit sarcomere shortening to propel thin filaments into the C-zone. PKA increased sarcomere shortening into the C-zone, while dephosphorylation of sMyBP-C by lambda phosphatase resulted in a deceleration of shortening and a brief recoil at sarcomere lengths near where the thin filaments should enter the C-zone implicating the potential for a substantive internal load that can oppose myofilament sliding. Beat-to-beat heart function is modulated by [beta]-adrenergic stimulation, which elicits PKA phosphorylation of several cardiac myocyte proteins including cardiac troponin I (cTnI). PKA phosphorylation of the cTnI is known to regulate myofilament Ca2+ sensitivity of force, augment cardiac myofilament power, transient force overshoot, and cross-bridge cycling kinetics. However, it is unknown if cTnI N-terminal phosphorylation per se affects myofilament power and stretch activation. We tested the hypothesis that cTnI N-terminal pseudo-phosphorylation will increase transient force overshoot, stretch activation, rates of force development, and power output consistent with a thin filament molecular mechanism regulating sarcomere function. Molecular sufficiency was investigated by measuring power and stretch activation after cTn exchange into rat permeabilized slow-twitch skeletal muscle fibers, since slow-skeletal TnI lacks N-terminal phosphorylation sites. cTnI pseudo-phosphorylation increased stretch activation and fiber power output generating capacity. Combinatorial S22/23D + Y26E cTnI synergistically increased rates of delayed force development. Taken together, these results suggest MyBP-C and its phosphorylation state regulates sarcomere contraction by modified cross-bridge cycling kinetics, cross-bridge recruitment, and altered internal drag forces that tend to oppose force generation, myofilament sliding, and power output. TnI and its phosphorylation state appears to regulate stretch activation and power output by a combination of redundant and synergistic biophysical mechanisms, which may complement each other to tune beat-to-beat myocardial output demand.

show abstract

Pathogenesis associated with a restrictive cardiomyopathy mutant in cardiac troponin T is due to reduced protein stability and greatly increased myofilament Ca2+ sensitivity

Cited by 7 publications

References 61 publications

Overexpression of miRNA-9 Generates Muscle Hypercontraction Through Translational Repression of Troponin-T inDrosophila melanogasterIndirect Flight Muscles

Overexpression of miRNA-9 Generates Muscle Hypercontraction Through Translational Repression of Troponin-T inDrosophila melanogasterIndirect Flight Muscles

Cardiomyocyte PKA Ablation Enhances Basal Contractility While Eliminates Cardiac β-Adrenergic Response Without Adverse Effects on the Heart

Molecular regulation of sarcomere function

Contact Info

Product

Resources

About