The Tropomyosin Binding Region of Cardiac Troponin T Modulates Crossbridge Recruitment Dynamics in Rat Cardiac Muscle Fibers

How different mutations in cardiac troponin T (cTnT) lead to distinct secondary downstream cellular remodeling in familial hypertrophic cardiomyopathy (FHC) remains elusive. To explore the molecular basis for the distinct impact of different mutations in cTnT on cardiac myocytes, we studied mechanical activity of detergent-skinned muscle fiber bundles from different lines of transgenic (TG) mouse hearts that express wild-type cTnT (WTTG), R92W cTnT, R92L cTnT, and Delta-160 cTnT (deletion of amino acid 160). The amount of mutant cTnT is ∼50% of the total myocellular cTnT in both R92W and R92L TG mouse hearts and ∼35% in Delta-160 TG mouse hearts. Myofilament Ca2+ sensitivity was enhanced in all mutant cTnT TG cardiac muscle fibers. Compared with the WTTG fibers, Ca2+ sensitivity increased significantly at short sarcomere length (SL) of 1.9 μm ( P < 0.001) in R92W TG fibers by 2.2-fold, in R92L by 2.0-fold, and in Delta-160 by 1.3-fold. At long SL of 2.3 μm, Ca2+ sensitivity increased significantly ( P < 0.01) in a similar manner (R92W, 2.5-fold; R92L, 1.9-fold; Delta-160, 1.3-fold). Ca2+-activated maximal tension remained unaltered in all TG muscle fibers. However, tension-dependent ATP consumption increased significantly in Delta-160 TG muscle fibers at both short SL (23%, P < 0.005) and long SL (37%, P < 0.0001), suggesting a mutation-induced change in cross-bridge detachment rate constant. Chronic stresses on relative cellular ATP level in cardiac myocytes may cause a strain on energy-dependent Ca2+ homeostatic mechanisms. This may result in pathological remodeling that we observed in Delta-160 TG cardiac myocytes where the ratio of sarco(endo)plasmic reticulum Ca2+-ATPase 2/phospholamban decreased significantly. Our results suggest that different types of stresses imposed on cardiac myocytes would trigger distinct cellular signaling, which leads to remodeling that may be unique to some mutants.

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Disease-Causing Mutations in Cardiac Troponin T: Identification of a Critical Tropomyosin-Binding Region

Biophysical Journal ◽

10.1016/s0006-3495(01)75924-3 ◽

2001 ◽

Vol 81 (5) ◽

pp. 2827-2837 ◽

Cited By ~ 102

Author(s):

Thomas Palm ◽

Sarah Graboski ◽

Sarah E. Hitchcock-DeGregori ◽

Norma J. Greenfield

Keyword(s):

Cardiac Troponin ◽

Troponin T ◽

Cardiac Troponin T ◽

Binding Region

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Functional effects of troponin T mutations found in hypertrophic cardiomyopathy on the Ca2+-activated contraction of skinned cardiac muscle fibers

The Japanese Journal of Pharmacology ◽

10.1016/s0021-5198(19)47931-6 ◽

2000 ◽

Vol 82 ◽

pp. 116

Author(s):

Sachio Morimoto ◽

Keita Harada ◽

Fumi Takahashi ◽

Reiko Minakami ◽

Iwao Ohtsuki

Keyword(s):

Hypertrophic Cardiomyopathy ◽

Cardiac Muscle ◽

Troponin T ◽

Muscle Fibers ◽

Cardiac Muscle Fibers

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Hypertrophic cardiomyopathy mutation in cardiac troponin T (R95H) attenuates length-dependent activation in guinea pig cardiac muscle fibers

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00369.2017 ◽

2017 ◽

Vol 313 (6) ◽

pp. H1180-H1189 ◽

Cited By ~ 6

Author(s):

Alexis V. Mickelson ◽

Murali Chandra

Keyword(s):

Hypertrophic Cardiomyopathy ◽

Guinea Pig ◽

Central Region ◽

Cardiac Troponin ◽

Sarcomere Length ◽

Troponin T ◽

Cardiac Troponin T ◽

Cross Bridge ◽

Cardiac Muscle Fibers ◽

Length Dependent Activation

The central region of cardiac troponin T (TnT) is important for modulating the dynamics of muscle length-mediated cross-bridge recruitment. Therefore, hypertrophic cardiomyopathy mutations in the central region may affect cross-bridge recruitment dynamics to alter myofilament Ca2+ sensitivity and length-dependent activation of cardiac myofilaments. Given the importance of the central region of TnT for cardiac contractile dynamics, we studied if hypertrophic cardiomyopathy-linked mutation (TnTR94H)-induced effects on contractile function would be differently modulated by sarcomere length (SL). Recombinant wild-type TnT (TnTWT) and the guinea pig analog of the human R94H mutation (TnTR95H) were reconstituted into detergent-skinned cardiac muscle fibers from guinea pigs. Steady-state and dynamic contractile measurements were made at short and long SLs (1.9 and 2.3 µm, respectively). Our results demonstrated that TnTR95H increased pCa50 (−log of free Ca2+ concentration) to a greater extent at short SL; TnTR95H increased pCa50 by 0.11 pCa units at short SL and 0.07 pCa units at long SL. The increase in pCa50 associated with an increase in SL from 1.9 to 2.3 µm (ΔpCa50) was attenuated nearly twofold in TnTR95H fibers; ΔpCa50 was 0.09 pCa units for TnTWT fibers but only 0.05 pCa units for TnTR95H fibers. The SL dependency of rate constants of cross-bridge distortion dynamics and tension redevelopment was also blunted by TnTR95H. Collectively, our observations on the SL dependency of pCa50 and rate constants of cross-bridge distortion dynamics and tension redevelopment suggest that mechanisms underlying the length-dependent activation cardiac myofilaments are attenuated by TnTR95H. NEW & NOTEWORTHY Mutant cardiac troponin T (TnTR95H) differently affects myofilament Ca2+ sensitivity at short and long sarcomere length, indicating that mechanisms underlying length-dependent activation are altered by TnTR95H. TnTR95H enhances myofilament Ca2+ sensitivity to a greater extent at short sarcomere length, thus attenuating the length-dependent increase in myofilament Ca2+ sensitivity.

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The Effect of HCM and RCM Causing Mutations of Troponin T on Force Generation and Cross-Bridge Kinetics in Thin-Filament Reconstituted Bovine Cardiac Muscle Fibers

Biophysical Journal ◽

10.1016/j.bpj.2011.11.853 ◽

2012 ◽

Vol 102 (3) ◽

pp. 156a

Author(s):

Fan Bai ◽

Hannah M. Caster ◽

Jose R. Pinto ◽

James D. Potter ◽

Masataka Kawai

Keyword(s):

Cardiac Muscle ◽

Thin Filament ◽

Troponin T ◽

Muscle Fibers ◽

Force Generation ◽

Cross Bridge ◽

Cardiac Muscle Fibers

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Crossbridge Recruitment Dynamics are Divergently Affected by α/β-Myosin Heavy Chain Isoforms in Cardiac Muscle Fibers Containing the Hypertrophic Cardiomyopathy Mutation (A30V) and Protein Kinase C Phosphomimetic (T203E) in Mouse Cardiac Troponin T

Biophysical Journal ◽

10.1016/j.bpj.2015.11.1590 ◽

2016 ◽

Vol 110 (3) ◽

pp. 295a

Author(s):

Alexis Mickelson ◽

Sampath Gollapudi ◽

Murali Chandra

Keyword(s):

Protein Kinase C ◽

Hypertrophic Cardiomyopathy ◽

Protein Kinase ◽

Heavy Chain ◽

Cardiac Troponin ◽

Troponin T ◽

Myosin Heavy Chain Isoforms ◽

Cardiac Troponin T ◽

Kinase C ◽

Cardiac Muscle Fibers

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Coexistence of cardiac troponin T variants reduces heart efficiency

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.01105.2009 ◽

2010 ◽

Vol 299 (1) ◽

pp. H97-H105 ◽

Cited By ~ 18

Author(s):

Han-Zhong Feng ◽

J.-P. Jin

Keyword(s):

Cardiac Muscle ◽

Transgenic Mouse ◽

Cardiac Troponin ◽

Troponin T ◽

Ex Vivo ◽

Heart Function ◽

Left Ventricular Pressure ◽

Left Ventricular ◽

Cardiac Troponin T ◽

Myofilament Activation

Corresponding to the synchronized contraction of the myocardium and rhythmic pumping function of the heart, a single form of cardiac troponin T (cTnT) is present in the adult cardiac muscle of humans and most other vertebrate species. Alternative splicing variants of cTnT are found in failing human hearts and animal dilated cardiomyopathies. Biochemical analyses have shown that these cTnT variants are functional and produce shifted myofilament Ca2+ sensitivity. We proposed a hypothesis that the coexistence of two or more functionally distinct TnT variants in the adult ventricular muscle that is normally activated as a syncytium may decrease heart function and cause cardiomyopathy (Huang et al., Am J Physiol Cell Physiol 294: C213–C222, 2008). In the present study, we studied transgenic mouse hearts expressing one or two cTnT variants in addition to normal adult cTnT to investigate whether desynchronized myofilament activation decreases ventricular efficiency. The function of ex vivo working hearts was examined in the absence of systemic neurohumoral influence. The results showed that the transgenic mouse hearts produced lower maximum left ventricular pressure, slower contractile and relaxation velocities, and decreased stroke volume compared with wild-type controls. Ventricular pumping efficiency, calculated by the ejection integral versus total systolic integral and cardiac work versus oxygen consumption, was significantly lower in transgenic mouse hearts and corresponded to the number of cTnT variants present. The results indicated a pathogenic mechanism in which the coexistence of functionally different cTnT variants in cardiac muscle reduces myocardial efficiency due to desynchronized thin filament activation.

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