Familial hypertrophic cardiomyopathy (FHC) is one of the pathological compensatory manifestations found in the heart resulting from its inability to adequately pump blood, thus leading to hypertrophy and often to premature cardiac death. Over the past 4 years our laboratory has been studying the functional consequences of several FHC mutations in the regulatory light chain (RLC) of myosin expressed in transgenic mice. We hypothesize that by changing the properties of the RLC Ca2+-Mg2+ binding site, the FHC mutations interfere with the intracellular function of RLC as a temporary delayed Ca2+- buffer and lead to increased or decreased kinetics of muscle relaxation. Another hypothesis pertains to the mutation controlled metal occupancy of the Ca2+-Mg2+ binding site of RLC and the mechanism by which Ca2+ or Mg2+ binding to RLC may influence the interaction of myosin with actin and tension generation. We further hypothesize that an FHC induced pathological cardiac phenotype can be rescued by Ca2+-calmodulin activated MLCK phosphorylation of the RLC-mutated myocardium. This application will continue the use of integrated multidisciplinary approaches from single molecule, cell to organ levels and novel transgenic mouse models to address the following questions:
SPECIFIC AIM 1 : Do FHC induced changes in the properties of the RLC Ca2+-Mg2+ binding site inhibit or facilitate the function of RLC as a temporary intracellular calcium buffer? Do FHC mutations shift the metal occupancy of the RLC Ca2+-Mg2+ binding site during muscle contraction? SPECIFIC AIM 2: Is RLC phosphorylation by Ca2+-calmodulin (CaM) activated myosin light chain kinase (MLCK) affected by FHC-linked RLC mutations? Can MLCK phosphorylation rescue a mutation induced pathological cardiac phenotype? SPECIFIC AIM 3: Do FHC-associated mutations in RLC alter intermolecular interactions between RLC and myosin heavy chain (HC) and ultimately myosin and actin? Do these changes lead to myofilament disarray, cardiac hypertrophy and dysfunction of the mutated myocardium? Successful execution of this proposal will result in new mechanical, physiological and histological information regarding the role of the RLC in cardiac muscle contraction in health and disease. Relevance: Cardiovascular diseases are the number one cause of mortality worldwide with heart failure being highly prevalent in most affluent parts of the world. There is an urgent need for a better understanding of the mechanisms underlying Familial Hypertrophic Cardiomyopathy (FHC) that often leads to premature sudden cardiac death (SCD). This proposal addresses the mechanisms by which the mutations in myosin regulatory light chain (RLC) cause FHC and lead to SCD. Determining the mechanisms of the RLC- mediated regulation of contraction in the healthy and hypertrophic heart will provide insight and be instrumental in developing specific therapeutic strategies that can be employed to reverse or prevent FHC RLC pathology.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL071778-09
Application #
8242787
Study Section
Special Emphasis Panel (ZRG1-CVS-P (03))
Program Officer
Adhikari, Bishow B
Project Start
2002-12-01
Project End
2014-03-31
Budget Start
2012-04-01
Budget End
2014-03-31
Support Year
9
Fiscal Year
2012
Total Cost
$383,943
Indirect Cost
$104,308
Name
University of Miami School of Medicine
Department
Pharmacology
Type
Schools of Medicine
DUNS #
052780918
City
Coral Gables
State
FL
Country
United States
Zip Code
33146
Muthu, Priya; Liang, Jingsheng; Schmidt, William et al. (2014) In vitro rescue study of a malignant familial hypertrophic cardiomyopathy phenotype by pseudo-phosphorylation of myosin regulatory light chain. Arch Biochem Biophys 552-553:29-39
Huang, Wenrui; Liang, Jingsheng; Kazmierczak, Katarzyna et al. (2014) Hypertrophic cardiomyopathy associated Lys104Glu mutation in the myosin regulatory light chain causes diastolic disturbance in mice. J Mol Cell Cardiol 74:318-29
Kazmierczak, Katarzyna; Paulino, Ellena C; Huang, Wenrui et al. (2013) Discrete effects of A57G-myosin essential light chain mutation associated with familial hypertrophic cardiomyopathy. Am J Physiol Heart Circ Physiol 305:H575-89
Michael, John Jeshurun; Gollapudi, Sampath K; Ford, Steven J et al. (2013) Deletion of 1-43 amino acids in cardiac myosin essential light chain blunts length dependency of Ca(2+) sensitivity and cross-bridge detachment kinetics. Am J Physiol Heart Circ Physiol 304:H253-9
Wang, Li; Muthu, Priya; Szczesna-Cordary, Danuta et al. (2013) Diversity and similarity of motor function and cross-bridge kinetics in papillary muscles of transgenic mice carrying myosin regulatory light chain mutations D166V and R58Q. J Mol Cell Cardiol 62:153-63
Kazmierczak, Katarzyna; Muthu, Priya; Huang, Wenrui et al. (2012) Myosin regulatory light chain mutation found in hypertrophic cardiomyopathy patients increases isometric force production in transgenic mice. Biochem J 442:95-103
Lu, Xiaoqing; Kazmierczak, Katarzyna; Jiang, Xiaoyu et al. (2011) Germinal center-specific protein human germinal center associated lymphoma directly interacts with both myosin and actin and increases the binding of myosin to actin. FEBS J 278:1922-31
Polewicz, Dorota; Cadete, Virgilio J J; Doroszko, Adrian et al. (2011) Ischemia induced peroxynitrite dependent modifications of cardiomyocyte MLC1 increases its degradation by MMP-2 leading to contractile dysfunction. J Cell Mol Med 15:1136-47
Muthu, Priya; Wang, Li; Yuan, Chen-Ching et al. (2011) Structural and functional aspects of the myosin essential light chain in cardiac muscle contraction. FASEB J 25:4394-405
Mettikolla, P; Calander, N; Luchowski, R et al. (2011) Cross-bridge kinetics in myofibrils containing familial hypertrophic cardiomyopathy R58Q mutation in the regulatory light chain of myosin. J Theor Biol 284:71-81

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