Takeshi Aiba1, Federica Farinelli1, Geran Kostecki1, Geoffrey G Hesketh1, David Edwards1, Subrata Biswas1, Leslie Tung1, Gordon F Tomaselli2. 1. From the Division of Cardiology, Department of Internal Medicine, Johns Hopkins University School of Medicine, Baltimore, MD (T.A., F.F., D.E., S.B., G.F.T.); and Department of Biomedical Engineering (G.K., L.T.) and Department of Biological Chemistry (G.G.H.), Johns Hopkins University, Baltimore, MD. 2. From the Division of Cardiology, Department of Internal Medicine, Johns Hopkins University School of Medicine, Baltimore, MD (T.A., F.F., D.E., S.B., G.F.T.); and Department of Biomedical Engineering (G.K., L.T.) and Department of Biological Chemistry (G.G.H.), Johns Hopkins University, Baltimore, MD. gtomasel@jhmi.edu.
Abstract
BACKGROUND: The mechanisms of the electrocardiographic changes and arrhythmias in Brugada syndrome (BrS) remain controversial. Mutations in the sodium channel gene, SCN5A, and regulatory proteins that reduce or eliminate sodium current (INa) have been linked to BrS. We studied the properties of a BrS-associated SCN5A mutation in a protein kinase A (PKA) consensus phosphorylation site, R526H. METHODS AND RESULTS: In vitro PKA phosphorylation was detected in the I-II linker peptide of wild-type (WT) channels but not R526H or S528A (phosphorylation site) mutants. Cell surface expression of R526H and S528A channels was reduced compared with WT. Whole-cell INa through all channel variants revealed no significant differences in the steady-state activation, inactivation, and recovery from inactivation. Peak current densities of the mutants were significantly reduced compared with WT. Infection of 2D cultures of neonatal rat ventricular myocytes with WT and mutant channels increased conduction velocity compared with noninfected cells. PKA stimulation significantly increased peak INa and conduction velocity of WT but not mutant channels. Oxidant stress inhibits cardiac INa; WT and mutant INa decreases with the intracellular application of reduced nicotinamide adenine dinucleotide (NADH), an effect that is reversed by PKA stimulation in WT but not in R526H or S528A channels. CONCLUSIONS: We identified a family with BrS and an SCN5A mutation in a PKA consensus phosphorylation site. The BrS mutation R526H is associated with a reduction in the basal level of INa and a failure of PKA stimulation to augment the current that may contribute to the predisposition to arrhythmias in patients with BrS, independent of the precipitants.
BACKGROUND: The mechanisms of the electrocardiographic changes and arrhythmias in Brugada syndrome (BrS) remain controversial. Mutations in the sodium channel gene, SCN5A, and regulatory proteins that reduce or eliminate sodium current (INa) have been linked to BrS. We studied the properties of a BrS-associated SCN5A mutation in a protein kinase A (PKA) consensus phosphorylation site, R526H. METHODS AND RESULTS: In vitro PKA phosphorylation was detected in the I-II linker peptide of wild-type (WT) channels but not R526H or S528A (phosphorylation site) mutants. Cell surface expression of R526H and S528A channels was reduced compared with WT. Whole-cell INa through all channel variants revealed no significant differences in the steady-state activation, inactivation, and recovery from inactivation. Peak current densities of the mutants were significantly reduced compared with WT. Infection of 2D cultures of neonatal rat ventricular myocytes with WT and mutant channels increased conduction velocity compared with noninfected cells. PKA stimulation significantly increased peak INa and conduction velocity of WT but not mutant channels. Oxidant stress inhibits cardiac INa; WT and mutant INa decreases with the intracellular application of reduced nicotinamide adenine dinucleotide (NADH), an effect that is reversed by PKA stimulation in WT but not in R526H or S528A channels. CONCLUSIONS: We identified a family with BrS and an SCN5A mutation in a PKA consensus phosphorylation site. The BrS mutation R526H is associated with a reduction in the basal level of INa and a failure of PKA stimulation to augment the current that may contribute to the predisposition to arrhythmias in patients with BrS, independent of the precipitants.
Authors: Mark G Hoogendijk; Tobias Opthof; Pieter G Postema; Arthur A M Wilde; Jacques M T de Bakker; Ruben Coronel Journal: Circ Arrhythm Electrophysiol Date: 2010-06
Authors: Haifa Hallaq; Dao W Wang; Jennifer D Kunic; Alfred L George; K Sam Wells; Katherine T Murray Journal: Am J Physiol Heart Circ Physiol Date: 2011-11-18 Impact factor: 4.733
Authors: Ivan A Adzhubei; Steffen Schmidt; Leonid Peshkin; Vasily E Ramensky; Anna Gerasimova; Peer Bork; Alexey S Kondrashov; Shamil R Sunyaev Journal: Nat Methods Date: 2010-04 Impact factor: 28.547
Authors: Alexey V Zaitsev; Natalia S Torres; Keiko M Cawley; Amira D Sabry; Junco S Warren; Mark Warren Journal: Am J Physiol Heart Circ Physiol Date: 2019-03-15 Impact factor: 4.733
Authors: Hassan Musa; Crystal F Kline; Amy C Sturm; Nathaniel Murphy; Sara Adelman; Chaojian Wang; Haidun Yan; Benjamin L Johnson; Thomas A Csepe; Ahmet Kilic; Robert S D Higgins; Paul M L Janssen; Vadim V Fedorov; Raul Weiss; Christina Salazar; Thomas J Hund; Geoffrey S Pitt; Peter J Mohler Journal: Proc Natl Acad Sci U S A Date: 2015-09-21 Impact factor: 11.205
Authors: Andrew M Glazer; Yuko Wada; Bian Li; Ayesha Muhammad; Olivia R Kalash; Matthew J O'Neill; Tiffany Shields; Lynn Hall; Laura Short; Marcia A Blair; Brett M Kroncke; John A Capra; Dan M Roden Journal: Am J Hum Genet Date: 2020-06-12 Impact factor: 11.025
Authors: Daniel S Matasic; Jin-Young Yoon; Jared M McLendon; Haider Mehdi; Mark S Schmidt; Alexander M Greiner; Pravda Quinones; Gina M Morgan; Ryan L Boudreau; Kaikobad Irani; Charles Brenner; Barry London Journal: J Mol Cell Cardiol Date: 2020-03-21 Impact factor: 5.000