title: E258K HCM-causing mutation in cardiac MyBP-C reduces contractile force and accelerates twitch kinetics by disrupting the cMyBP-C and myosin S2 interaction.
abstract: Mutations in cardiac myosin binding protein C (cMyBP-C) are prevalent causes of hypertrophic cardiomyopathy (HCM). Although HCM-causing truncation mutations in cMyBP-C are well studied, the growing number of disease-related cMyBP-C missense mutations remain poorly understood. Our objective was to define the primary contractile effect and molecular disease mechanisms of the prevalent cMyBP-C E258K HCM-causing mutation in nonremodeled murine engineered cardiac tissue (mECT). Wild-type and human E258K cMyBP-C were expressed in mECT lacking endogenous mouse cMyBP-C through adenoviral-mediated gene transfer. Expression of E258K cMyBP-C did not affect cardiac cell survival and was appropriately incorporated into the cardiac sarcomere. Functionally, expression of E258K cMyBP-C caused accelerated contractile kinetics and severely compromised twitch force amplitude in mECT. Yeast two-hybrid analysis revealed that E258K cMyBP-C abolished interaction between the N terminal of cMyBP-C and myosin heavy chain sub-fragment 2 (S2). Furthermore, this mutation increased the affinity between the N terminal of cMyBP-C and actin. Assessment of phosphorylation of three serine residues in cMyBP-C showed that aberrant phosphorylation of cMyBP-C is unlikely to be responsible for altering these interactions. We show that the E258K mutation in cMyBP-C abolishes interaction between N-terminal cMyBP-C and myosin S2 by directly disrupting the cMyBP-C-S2 interface, independent of cMyBP-C phosphorylation. Similar to cMyBP-C ablation or phosphorylation, abolition of this inhibitory interaction accelerates contractile kinetics. Additionally, the E258K mutation impaired force production of mECT, which suggests that in addition to the loss of physiological function, this mutation disrupts contractility possibly by tethering the thick and thin filament or acting as an internal load.
authors: Department of Pediatrics, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA.
authors: Willem J
authors: WJ
authors: De Lange
authors: Adrian C
authors: AC
authors: Grimes
authors: Laura F
authors: LF
authors: Hegge
authors: Alexander M
authors: AM
authors: Spring
authors: Taylor M
authors: TM
authors: Brost
authors: J Carter
authors: JC
authors: Ralphe
mesh_terms: D000199:Actins / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002312:Cardiomyopathy, Hypertrophic / Q000235:genetics*; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D051379:Mice; D053580:Muscle Strength / Q000235:genetics*; D009154:Mutation; D009200:Myocardial Contraction / Q000235:genetics*; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D018995:Myosin Heavy Chains / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012518:Sarcomeres / Q000378:metabolism* / Q000502:physiology; D000099163:Myosin Binding Protein C
chemical_list: D000199:Actins; D002352:Carrier Proteins; D018995:Myosin Heavy Chains; D000099163:Myosin Binding Protein C
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