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Phosphomimetic‐mediated in vitro rescue of hypertrophic cardiomyopathy linked to R58Q mutation in myosin regulatory light chain
Author(s) -
Yadav Sunil,
Kazmierczak Katarzyna,
Liang Jingsheng,
Sitbon Yoel H.,
SzczesnaCordary Danuta
Publication year - 2019
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14702
Subject(s) - myosin , actin , myosin light chain kinase , phosphorylation , troponin , tropomyosin , myh7 , biology , myosin atpase , adenosine triphosphate , biochemistry , chemistry , microbiology and biotechnology , atpase , medicine , myocardial infarction , enzyme
Myosin regulatory light chain ( RLC ) phosphorylation is important for cardiac muscle mechanics/function as well as for the Ca 2+ ‐troponin/tropomyosin regulation of muscle contraction. This study focuses on the arginine to glutamine (R58Q) substitution in the human ventricular RLC ( MYL 2 gene), linked to malignant hypertrophic cardiomyopathy in humans and causing severe functional abnormalities in transgenic (Tg) R58Q mice, including inhibition of cardiac RLC phosphorylation. Using a phosphomimic recombinant RLC variant where Ser‐15 at the phosphorylation site was substituted with aspartic acid (S15D) and placed in the background of R58Q, we aimed to assess whether we could rescue/mitigate R58Q‐induced structural/functional abnormalities in vitro . We show rescue of several R58Q‐exerted adverse phenotypes in S15D‐R58Q‐reconstituted porcine cardiac muscle preparations. A low level of maximal isometric force observed for R58Q‐ versus WT ‐reconstituted fibers was restored by S15D‐R58Q. Significant beneficial effects were also observed on the V max of actin‐activated myosin ATP ase activity in S15D‐R58Q versus R58Q‐reconstituted myosin, along with its binding to fluorescently labeled actin. We also report that R58Q promotes the OFF state of myosin, both in reconstituted porcine fibers and in Tg mouse papillary muscles, thereby stabilizing the super‐relaxed state ( SRX ) of myosin, characterized by a very low ATP turnover rate. Experiments in S15D‐R58Q‐reconstituted porcine fibers showed a mild destabilization of the SRX state, suggesting an S15D‐mediated shift in disordered‐relaxed ( DRX )↔ SRX equilibrium toward the DRX state of myosin. Our study shows that S15D‐phosphomimic can be used as a potential rescue strategy to abrogate/alleviate the RLC mutation‐induced phenotypes and is a likely candidate for therapeutic intervention in HCM patients.