Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions

D. Duggal, J. Nagwekar, R. Rich, W. Huang, K. Midde, Rafal Fudala, Hriday Das, Ignacy Gryczynski, D. Szczesna-Cordary, Julian Borejdo

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4 Citations (Scopus)

Abstract

Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ~20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contractionrelaxation cycle may contribute to altered contractility and the development of FHC.

Original languageEnglish
Pages (from-to)H1248-H1257
JournalAmerican Journal of Physiology - Heart and Circulatory Physiology
Volume308
Issue number10
DOIs
StatePublished - 1 Jan 2015

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Myosin Light Chains
Myosins
Familial Hypertrophic Cardiomyopathy
Actins
Mutation
Myofibrils
Photobleaching
Sudden Cardiac Death
Mutation Rate
Transgenic Mice
Heart Ventricles
Coloring Agents
Adenosine Triphosphate
Stroke
Light

Keywords

  • Left ventricle
  • Mutation of regulatory light chain
  • Polarization of fluorescence

Cite this

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title = "Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions",
abstract = "Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ~20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contractionrelaxation cycle may contribute to altered contractility and the development of FHC.",
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Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions. / Duggal, D.; Nagwekar, J.; Rich, R.; Huang, W.; Midde, K.; Fudala, Rafal; Das, Hriday; Gryczynski, Ignacy; Szczesna-Cordary, D.; Borejdo, Julian.

In: American Journal of Physiology - Heart and Circulatory Physiology, Vol. 308, No. 10, 01.01.2015, p. H1248-H1257.

Research output: Contribution to journalArticleResearchpeer-review

TY - JOUR

T1 - Effect of a myosin regulatory light chain mutation K104E on actin-myosin interactions

AU - Duggal, D.

AU - Nagwekar, J.

AU - Rich, R.

AU - Huang, W.

AU - Midde, K.

AU - Fudala, Rafal

AU - Das, Hriday

AU - Gryczynski, Ignacy

AU - Szczesna-Cordary, D.

AU - Borejdo, Julian

PY - 2015/1/1

Y1 - 2015/1/1

N2 - Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ~20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contractionrelaxation cycle may contribute to altered contractility and the development of FHC.

AB - Familial hypertrophic cardiomyopathy (FHC) is the most common cause of sudden cardiac death in young individuals. Molecular mechanisms underlying this disorder are largely unknown; this study aims at revealing how disruptions in actin-myosin interactions can play a role in this disorder. Cross-bridge (XB) kinetics and the degree of order were examined in contracting myofibrils from the ex vivo left ventricles of transgenic (Tg) mice expressing FHC regulatory light chain (RLC) mutation K104E. Because the degree of order and the kinetics are best studied when an individual XB makes a significant contribution to the overall signal, the number of observed XBs in an ex vivo ventricle was minimized to ~20. Autofluorescence and photobleaching were minimized by labeling the myosin lever arm with a relatively long-lived red-emitting dye containing a chromophore system encapsulated in a cyclic macromolecule. Mutated XBs were significantly better ordered during steady-state contraction and during rigor, but the mutation had no effect on the degree of order in relaxed myofibrils. The K104E mutation increased the rate of XB binding to thin filaments and the rate of execution of the power stroke. The stopped-flow experiments revealed a significantly faster observed dissociation rate in Tg-K104E vs. Tg-wild-type (WT) myosin and a smaller second-order ATP-binding rate for the K104E compared with WT myosin. Collectively, our data indicate that the mutation-induced changes in the interaction of myosin with actin during the contractionrelaxation cycle may contribute to altered contractility and the development of FHC.

KW - Left ventricle

KW - Mutation of regulatory light chain

KW - Polarization of fluorescence

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U2 - 10.1152/ajpheart.00834.2014

DO - 10.1152/ajpheart.00834.2014

M3 - Article

VL - 308

SP - H1248-H1257

JO - American Journal of Physiology - Heart and Circulatory Physiology

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SN - 0363-6135

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