title: A hypertrophic cardiomyopathy-associated MYBPC3 mutation common in populations of South Asian descent causes contractile dysfunction.
abstract: Hypertrophic cardiomyopathy (HCM) results from mutations in genes encoding sarcomeric proteins, most often MYBPC3, which encodes cardiac myosin binding protein-C (cMyBP-C). A recently discovered HCM-associated 25-base pair deletion in MYBPC3 is inherited in millions worldwide. Although this mutation causes changes in the C10 domain of cMyBP-C (cMyBP-C(C10mut)), which binds to the light meromyosin (LMM) region of the myosin heavy chain, the underlying molecular mechanism causing HCM is unknown. In this study, adenoviral expression of cMyBP-C(C10mut) in cultured adult rat cardiomyocytes was used to investigate protein localization and evaluate contractile function and Ca(2+) transients, compared with wild-type cMyBP-C expression (cMyBP-C(WT)) and controls. Forty-eight hours after infection, 44% of cMyBP-C(WT) and 36% of cMyBP-C(C10mut) protein levels were determined in total lysates, confirming equal expression. Immunofluorescence experiments showed little or no localization of cMyBP-C(C10mut) to the C-zone, whereas cMyBP-C(WT) mostly showed C-zone staining, suggesting that cMyBP-C(C10mut) could not properly integrate in the C-zone of the sarcomere. Subcellular fractionation confirmed that most cMyBP-C(C10mut) resided in the soluble fraction, with reduced presence in the myofilament fraction. Also, cMyBP-C(C10mut) displayed significantly reduced fractional shortening, sarcomere shortening, and relaxation velocities, apparently caused by defects in sarcomere function, because Ca(2+) transients were unaffected. Co-sedimentation and protein cross-linking assays confirmed that C10(mut) causes the loss of C10 domain interaction with myosin LMM. Protein homology modeling studies showed significant structural perturbation in cMyBP-C(C10mut), providing a potential structural basis for the alteration in its mode of interaction with myosin LMM. Therefore, expression of cMyBP-C(C10mut) protein is sufficient to cause contractile dysfunction in vitro.
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153, and.
authors.forename: Diederik W D
authors.initials: DW
authors.lastname: Kuster
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153, and.
authors.forename: Suresh
authors.initials: S
authors.lastname: Govindan
authors.affiliation: the Department of Microbiology and.
authors.forename: Tzvia I
authors.initials: TI
authors.lastname: Springer
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153, and.
authors.forename: Jody L
authors.initials: JL
authors.lastname: Martin
authors.affiliation: the Department of Microbiology and the Cell, Molecular, and Structural Biology Program, Miami University, Oxford, Ohio 45056.
authors.forename: Natosha L
authors.initials: NL
authors.lastname: Finley
authors.affiliation: From the Department of Cell and Molecular Physiology, Health Sciences Division, Loyola University Chicago, Maywood, Illinois 60153, and ssadayappan@luc.edu.
authors.forename: Sakthivel
authors.initials: S
authors.lastname: Sadayappan
mesh_terms: D000818:Animals; D001208:Asia; D044466:Asian People / Q000235:genetics; D002312:Cardiomyopathy, Hypertrophic / Q000208:ethnology / Q000235:genetics* / Q000503:physiopathology; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D048429:Cell Size; D002478:Cells, Cultured; D020022:Genetic Predisposition to Disease / Q000208:ethnology / Q000235:genetics*; D006801:Humans; D015151:Immunoblotting; D008297:Male; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D009119:Muscle Contraction / Q000235:genetics; D009154:Mutation; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D015879:Myosin Subfragments / Q000378:metabolism; D011485:Protein Binding / Q000235:genetics; D017434:Protein Structure, Tertiary; D017207:Rats, Sprague-Dawley; D012518:Sarcomeres / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism; D000099163:Myosin Binding Protein C
chemical_list: D002352:Carrier Proteins; D015879:Myosin Subfragments; D000099163:Myosin Binding Protein C
keywords: Cardiomyopathy; Contractile Protein; Genetic Polymorphism; Heart Failure; Muscle Physiology