BMC Medical Genetics
Establishing disease causality for a novel gene variant in familial dilated cardiomyopathy using a functional in-vitro assay of regulated thin filaments and human cardiac myosin
Euan A. Ashley3  Kathleen M. Ruppel2  James A. Spudich4  Susan M. Miller1  Shirley Sutton4  Suman Nag4  Karim I. Sallam3  Ruth F. Sommese6  Stephen Pan5 
[1]Department of Pharmaceutical Chemistry, University of California San Francisco, San Francisco, CA, USA
[2]Department of Pediatrics (Cardiology), Stanford University School of Medicine, Stanford, CA, USA
[3]Departments of Medicine (Cardiovascular Medicine), Stanford University School of Medicine, Stanford, CA, USA
[4]Department of Biochemistry, Stanford University School of Medicine, Stanford, CA, USA
[5]Leon H. Charney Division of Cardiology, NYU Langone Medical Center, New York, NY, USA
[6]Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, MN, USA
关键词: Gene mutation;    Calcium;    Myosin;    Troponin T;    Cardiomyopathy;   
Others  :  1230635
DOI  :  10.1186/s12881-015-0243-5
 received in 2014-10-28, accepted in 2015-10-16, published in 16
【 摘 要 】


As next generation sequencing for the genetic diagnosis of cardiovascular disorders becomes more widely used, establishing causality for putative disease causing variants becomes increasingly relevant. Diseases of the cardiac sarcomere provide a particular challenge in this regard because of the complexity of assaying the effect of genetic variants in human cardiac contractile proteins.


In this study we identified a novel variant R205Q in the cardiac troponin T gene (TNNT2). Carriers of the variant allele exhibited increased chamber volumes associated with decreased left ventricular ejection fraction. To clarify the causal role of this variant, we generated recombinant variant human protein and examined its calcium kinetics as well as the maximally activated ADP release of human β-cardiac myosin with regulated thin filaments containing the mutant troponin T. We found that the R205Q mutation significantly decreased the calcium sensitivity of the thin filament by altering the effective calcium dissociation kinetics.


The development of moderate throughput post-genomic assays is an essential step in the realization of the potential of next generation sequencing. Although technically challenging, biochemical and functional assays of human cardiac contractile proteins of the thin filament can be achieved and provide an orthogonal source of information to inform the question of causality for individual variants.

【 授权许可】

2015 Pan et al.

【 预 览 】
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