title: Restrictive Cardiomyopathy Troponin I R145W Mutation Does Not Perturb Myofilament Length-dependent Activation in Human Cardiac Sarcomeres.
abstract: The cardiac troponin I (cTnI) R145W mutation is associated with restrictive cardiomyopathy (RCM). Recent evidence suggests that this mutation induces perturbed myofilament length-dependent activation (LDA) under conditions of maximal protein kinase A (PKA) stimulation. Some cardiac disease-causing mutations, however, have been associated with a blunted response to PKA-mediated phosphorylation; whether this includes LDA is unknown. Endogenous troponin was exchanged in isolated skinned human myocardium for recombinant troponin containing either cTnI R145W, PKA/PKC phosphomimetic charge mutations (S23D/S24D and T143E), or various combinations thereof. Myofilament Ca2+sensitivity of force, tension cost, LDA, and single myofibril activation/relaxation parameters were measured. Our results show that both R145W and T143E uncouple the impact of S23D/S24D phosphomimetic on myofilament function, including LDA. Molecular dynamics simulations revealed a marked reduction in interactions between helix C of cTnC (residues 56, 59, and 63), and cTnI (residue 145) in the presence of either cTnI RCM mutation or cTnI PKC phosphomimetic. These results suggest that the RCM-associated cTnI R145W mutation induces a permanent structural state that is similar to, but more extensive than, that induced by PKC-mediated phosphorylation of cTnI Thr-143. We suggest that this structural conformational change induces an increase in myofilament Ca2+sensitivity and, moreover, uncoupling from the impact of phosphorylation of cTnI mediated by PKA at the Ser-23/Ser-24 target sites. The R145W RCM mutation by itself, however, does not impact LDA. These perturbed biophysical and biochemical myofilament properties are likely to significantly contribute to the diastolic cardiac pump dysfunction that is seen in patients suffering from a restrictive cardiomyopathy that is associated with the cTnI R145W mutation.
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153.
authors.forename: Alexey V
authors.initials: AV
authors.lastname: Dvornikov
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153.
authors.forename: Nikolai
authors.initials: N
authors.lastname: Smolin
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153.
authors.forename: Mengjie
authors.initials: M
authors.lastname: Zhang
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153.
authors.forename: Jody L
authors.initials: JL
authors.lastname: Martin
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153.
authors.forename: Seth L
authors.initials: SL
authors.lastname: Robia
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153 pdetombe@luc.edu.
authors.forename: Pieter P
authors.initials: PP
authors.lastname: de Tombe
mesh_terms: D019943:Amino Acid Substitution; D002313:Cardiomyopathy, Restrictive / Q000235:genetics / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D056004:Molecular Dynamics Simulation; D020125:Mutation, Missense; D011493:Protein Kinase C / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012518:Sarcomeres / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship; D019210:Troponin I / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D019210:Troponin I; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C
keywords: contractile protein; contractility; heart; length-dependent activation; molecular dynamics; myocardium; myofilament; phosphorylation; troponin