Searched for: person:coetzw01
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A homozygous SCN5A mutation associated with atrial standstill and sudden death
Tan, Reina Bianca; Gando, Ivan; Bu, Lei; Cecchin, Frank; Coetzee, William
BACKGROUND:Atrial standstill is an arrhythmogenic condition characterized by the absence of spontaneous electrical and mechanical atrial activity or in response to stimulation. There are few reported familial cases which have been associated with SCN5A mutations co-segregating with GJA5 or RYR2 however isolated SCN5A mutations are rare. OBJECTIVE:The purpose of this study was to determine the clinical and biophysical consequence of a novel SCN5A mutation identified in a family with progressive atrial standstill and sudden death. METHODS:The family of a sporadic case of congenital atrial standstill underwent genetic screening. Human Embryonic Kidney 293 cells were transfected with wild-type (WT) or mutant SCN5A cDNAs. Biophysical properties were studied using whole-cell using patch clamp methods. RESULTS:A novel homozygous SCN5A mutation, p.V1340L was identified in the proband and her sister. The proband had complete atrial standstill whereas the sister had partial atrial standstill. Heterozygous mutations were identified in the mother, father and brother. All three had normal sinus rhythm and were asymptomatic. The mutant Nav1.5(V1340L) reduced Nav1.5 current density as well as showed a depolarizing shift in the voltage-dependent steady-state activation (WT: -35.3±1.62 mV; V1340L: -22.4±2.59 mV; P = 0.001). CONCLUSIONS:A homozygous loss-of-function SCN5A mutation likely results in atrial standstill and sudden death due to suppression of initiation of action potential.
PMID: 29781517
ISSN: 1540-8159
CID: 3129702
Isoproterenol-induced action potential shortening mediated by sur1-containing KATP channels in human ips-derived atrial cardiomyocytes [Meeting Abstract]
Lader, J M; Lin, B; Yang, H; Coetzee, W A; Bu, L; Gelb, B D; Fishman, G I
Background: KAT P channels couple cellular metabolism and electrophysiology. Their molecular composition varies in different tissues and species. Rodent atrial KAT P channels have the SUR1 regulatory subunit, are activated by diazoxide and have been implicated in arrhythmogenesis in hypertension and excess beta-adrenergic tone. In contrast, human atrial KATP channels are insensitive to diazoxide and modulate APD only during extreme metabolic stress, where the SUR2A regulatory subunit is thought to be predominant. Objective: We hypothesized that changes in the human atrial action potential associated with beta-agonism are mediated by recruitment of SUR1-containing KATP channels. Methods: We used human induced pluripotent stem cell (hiPSC)-derived atrial cardiomyocytes where expression of a fuorescent reporter is driven by the atrial-specifc gene sarcolipin. Atrial specifcation was induced with retinoic acid. Di-4-ANBDQBS was used to perform optical action potential measurements on days 65-80 of differentiation. Excised patch clamping was used to evaluate KAT P channel density. Heterozygous ABCC8 (SUR1+/-) cells were generated using CRISPR/CAS9. Results: Optical mapping data are for APD90 with stimulation at 1.25 Hz The combination of isoproterenol (ISO, 10mu M) and rolipram (ROL, 10mu M) abbreviated APD compared to control (247.4+/-12.5ms, n=16 vs 344.2+/-22.9ms, n=22; p=0.002). This was ameliorated by 10mu M glibenclamide (312.0+/-18.9ms, n=23 vs 247.4+/-12.5ms, n=16; p=0.01). More patches from cells exposed to ISO and ROL had functional KATP channels (4/22 vs 0/24, p=0.045). Diazoxide shortened APD (267.3+/-21.7ms, n=20 vs 344.2+/-22.9ms, n=22; p=0.02). This was potentiated by prior beta-agonism (179.7+/-14.3ms, n=18 vs 267.3+/-21.7ms, n=20; p=0.002). Deletion of one ABCC8 allele ameliorated APD shortening with exposure to ISO, ROL, and diazoxide (240.9+/-18.2ms, n=14 vs 179.7+/-14.3ms, n=18; p=0.012). Functional KATP channel density after exposure to beta-agonists was reduced in SUR1+/-cells (1/40 vs 4/22, p=0.049). Conclusion: SUR1-containing KATP channels partially mediate beta-adrenergic APD shortening in human atrial cells and may represent a therapeutic target for atrial arrhythmia prevention
EMBASE:622469922
ISSN: 1556-3871
CID: 3151332
Regulation of KATPChannel Trafficking in Pancreatic β Cells by Protein Histidine Phosphorylation
Srivastava, Shekhar; Li, Zhai; Soomro, Irfana; Sun, Ying; Wang, Jianhui; Bao, Li; Coetzee, William A; Stanley, Charles A; Li, Chonghong; Skolnik, Edward Y
Protein histidine phosphatase 1 (PHPT-1) is an evolutionarily conserved 14 kDa protein that dephosphorylates phosphohistidine.PHPT-1
PMCID:5909995
PMID: 29440278
ISSN: 1939-327x
CID: 2958302
The trafficking protein, EHD2, positively regulates cardiac sarcolemmal KATP channel surface expression: role in cardioprotection
Yang, Hua Qian; Jana, Kundan; Rindler, Michael J; Coetzee, William A
ATP-sensitive K+ (KATP) channels uniquely link cellular energy metabolism to membrane excitability and are expressed in diverse cell types that range from the endocrine pancreas to neurons and smooth, skeletal, and cardiac muscle. A decrease in the surface expression of KATP channels has been linked to various disorders, including dysregulated insulin secretion, abnormal blood pressure, and impaired resistance to cardiac injury. In contrast, up-regulation of KATP channel surface expression may be protective, for example, by mediating the beneficial effect of ischemic preconditioning. Molecular mechanisms that regulate KATP channel trafficking are poorly understood. Here, we used cellular assays with immunofluorescence, surface biotinylation, and patch clamping to demonstrate that Eps15 homology domain-containing protein 2 (EHD2) is a novel positive regulator of KATP channel trafficking to increase surface KATP channel density. EHD2 had no effect on cardiac Na+ channels (Nav1.5). The effect is specific to EHD2 as other members of the EHD family-EHD1, EHD3, and EHD4-had no effect on KATP channel surface expression. EHD2 did not directly affect KATP channel properties as unitary conductance and ATP sensitivity were unchanged. Instead, we observed that the mechanism by which EHD2 increases surface expression is by stabilizing KATP channel-containing caveolar structures, which results in a reduced rate of endocytosis. EHD2 also regulated KATP channel trafficking in isolated cardiomyocytes, which validated the physiologic relevance of these observations. Pathophysiologically, EHD2 may be cardioprotective as a dominant-negative EHD2 mutant sensitized cardiomyocytes to ischemic damage. Our findings highlight EHD2 as a potential pharmacologic target in the treatment of diseases with KATP channel trafficking defects.-Yang, H. Q., Jana, K., Rindler, M. J., Coetzee, W. A. The trafficking protein, EHD2, positively regulates cardiac sarcolemmal KATP channel surface expression: role in cardioprotection.
PMCID:5892718
PMID: 29133341
ISSN: 1530-6860
CID: 2785362
Population-Based Mathematical Modeling to Deduce Disease-Causing Cardiac Na+ Channel Gating Defects [Meeting Abstract]
Campana, Chiara; Gando, Ivan; Tan, Reina Bianca; Cecchin, Frank; Coetzee, William A.; Sobie, Eric A.
ISI:000430563300167
ISSN: 0006-3495
CID: 3084792
Applying High-Resolution Variant Classification to Cardiac Arrhythmogenic Gene Testing in a Demographically Diverse Cohort of Sudden Unexplained Deaths
Lin, Ying; Williams, Nori; Wang, Dawei; Coetzee, William; Zhou, Bo; Eng, Lucy S; Um, Sung Yon; Bao, Ruijun; Devinsky, Orrin; McDonald, Thomas V; Sampson, Barbara A; Tang, Yingying
BACKGROUND:Genetic variant interpretation contributes to testing yield differences reported for sudden unexplained death. Adapting a high-resolution variant interpretation framework, which considers disease prevalence, reduced penetrance, genetic heterogeneity, and allelic contribution to determine the maximum tolerated allele count in gnomAD, we report an evaluation of cardiac channelopathy and cardiomyopathy genes in a large, demographically diverse sudden unexplained death cohort that underwent thorough investigation in the United States' largest medical examiner's office. METHODS AND RESULTS/RESULTS:The cohort has 296 decedents: 147 Blacks, 64 Hispanics, 49 Whites, 22 Asians, and 14 mixed ethnicities; 142 infants (1 to 11 months), 39 children (1 to 17 years), 74 young adults (18 to 34 years), and 41 adults (35 to 55 years). Eighty-nine cardiac disease genes were evaluated. Using a high-resolution variant interpretation workflow, we classified 17 variants as pathogenic or likely pathogenic (2 of which were incidental findings and excluded in testing yield analysis), 46 novel variants of uncertain significance, and 130 variants of uncertain significance. Nine pathogenic or likely pathogenic variants in ClinVar were reclassified to likely benign and excluded in testing yield analysis. The yields of positive cases by ethnicity and age were 21.4% in mixed ethnicities, 10.2% Whites, 4.5% Asians, 3.1% Hispanics, and 2% Blacks; 7.7% children, 7.3% in adults, 5.4% young adults, and 2.8% infants. The percentages of uncertain cases with variants of uncertain significance by ethnicity were 45.5% in Asians, 45.3% Hispanics, 44.20% Blacks, 36.7% Whites, and 14.3% in mixed ethnicities. CONCLUSIONS:High-resolution variant interpretation provides diagnostic accuracy and healthcare efficiency. Under-represented populations warrant greater inclusion in future studies.
PMID: 29247119
ISSN: 1942-3268
CID: 2892682
Plakophilin-2 is required for transcription of genes that control calcium cycling and cardiac rhythm
Cerrone, Marina; Montnach, Jerome; Lin, Xianming; Zhao, Yan-Ting; Zhang, Mingliang; Agullo-Pascual, Esperanza; Leo-Macias, Alejandra; Alvarado, Francisco J; Dolgalev, Igor; Karathanos, Thomas V; Malkani, Kabir; Van Opbergen, Chantal J M; van Bavel, Joanne J A; Yang, Hua-Qian; Vasquez, Carolina; Tester, David; Fowler, Steven; Liang, Fengxia; Rothenberg, Eli; Heguy, Adriana; Morley, Gregory E; Coetzee, William A; Trayanova, Natalia A; Ackerman, Michael J; van Veen, Toon A B; Valdivia, Hector H; Delmar, Mario
Plakophilin-2 (PKP2) is a component of the desmosome and known for its role in cell-cell adhesion. Mutations in human PKP2 associate with a life-threatening arrhythmogenic cardiomyopathy, often of right ventricular predominance. Here, we use a range of state-of-the-art methods and a cardiomyocyte-specific, tamoxifen-activated, PKP2 knockout mouse to demonstrate that in addition to its role in cell adhesion, PKP2 is necessary to maintain transcription of genes that control intracellular calcium cycling. Lack of PKP2 reduces expression of Ryr2 (coding for Ryanodine Receptor 2), Ank2 (coding for Ankyrin-B), Cacna1c (coding for CaV1.2) and Trdn (coding for triadin), and protein levels of calsequestrin-2 (Casq2). These factors combined lead to disruption of intracellular calcium homeostasis and isoproterenol-induced arrhythmias that are prevented by flecainide treatment. We propose a previously unrecognized arrhythmogenic mechanism related to PKP2 expression and suggest that mutations in PKP2 in humans may cause life-threatening arrhythmias even in the absence of structural disease.It is believed that mutations in desmosomal adhesion complex protein plakophilin 2 (PKP2) cause arrhythmia due to loss of cell-cell communication. Here the authors show that PKP2 controls the expression of proteins involved in calcium cycling in adult mouse hearts, and that lack of PKP2 can cause arrhythmia in a structurally normal heart.
PMCID:5524637
PMID: 28740174
ISSN: 2041-1723
CID: 2653852
Infant sudden death: mutations responsible for impaired Nav1.5 channel trafficking and function
Gando, Ivan; Morganstein, Jace; Jana, Kundan; McDonald, Thomas V; Tang, Yingying; Coetzee, William A
BACKGROUND: Two genetic variants in SCN5A, encoding the Nav1.5 Na+ channel alpha-subunit, were found in a five month-old girl who died suddenly in her sleep. The first variant is a missense mutation, resulting in an amino acid change (Q1832E), which has been described (but not characterized) in a patient with Brugada syndrome. The second is a nonsense mutation that produces a premature stop codon and a C-terminal truncation (R1944Delta). METHODS AND RESULTS: To investigate their functional relevance with patch clamp experiments in transfected HEK293 cells. The Q1832E mutation drastically reduced Nav1.5 current density. The R1944Delta C-terminal truncation had negligible effects on Nav1.5 current density. Neither of the mutations affected the voltage dependence of steady activation and inactivation or influenced the late Na+ current or the recovery from inactivation. Biochemical and immunofluorescent approaches demonstrated that the Q1832E mutation caused severe trafficking defects. PCR cloning and sequencing the victim's genomic DNA allowed us to determine that the two variants were in trans. We investigated the functional consequences by co-expressing Nav1.5(Q1832E) and Nav1.5(R1944Delta), which led to a significantly reduced current amplitude relative to wild-type. CONCLUSIONS: These SIDS-related variants caused a severely dysfunctional Nav1.5 channel, which was mainly due to trafficking defects caused by the Q1832E mutation. The decreased current density is likely to be a major contributing factor to arrhythmogenesis in Brugada syndrome and the sudden death of this SIDS victim
PMID: 28370132
ISSN: 1540-8159
CID: 2521352
Plasticity of sarcolemmal KATP channel surface expression during ischemia and ischemic preconditioning
Yang, Hua-Qian; Foster, Monique N; Jana, Kundan; Ho, Joanne; Rindler, Michael J; Coetzee, William A
AIMS: Myocardial ischemia remains the prime cause of morbidity and mortality in the United States. Ischemic preconditioning (IPC) is a powerful form of endogenous protection against myocardial infarction. We studied alterations in KATPchannels surface density as a potential mechanism of IPC's protection. METHODS AND RESULTS: Using cardiac-specific knockout of Kir6.2 subunits, we demonstrate an essential role for sarcolemmal KATPchannels in the infarct-limiting effect of IPC in the mouse heart. With biochemical membrane fractionation, we demonstrated that sarcolemmal KATPchannel subunits are distributed both to the sarcolemma and intracellular endosomal compartments. Global ischemia causes a loss of sarcolemmal KATPchannel subunit distribution and internalization to endosomal compartments. Ischemia-induced internalization of KATPchannels was prevented by CaMKII inhibition. KATPchannel subcellular redistribution was also observed with immunohistochemistry. Ischemic preconditioning prior to the index ischemia reduces not only the infarct size, but also prevents KATPchannel internalization. Furthermore, not only did adenosine mimic IPC by preventing infarct size, but it also prevented ischemia-induced KATPchannel internalization via a PKC-mediated pathway. We show that preventing endocytosis with dynasore reduces both KATPchannel internalization and strongly mitigates infarct development. CONCLUSIONS: Our data demonstrate that plasticity of KATPchannel surface expression must be considered as a potentially important mechanism of the protective effects of IPC and adenosine.
PMCID:4935516
PMID: 27037371
ISSN: 1522-1539
CID: 2059432
Disruption of ATP-sensitive potassium channel function in skeletal muscles promotes production and secretion of musclin Condensed title: Skeletal muscle I regulates musclin
Sierra, Ana; Subbotina, Ekaterina; Zhu, Zhiyong; Gao, Zhan; Koganti, Siva Rama Krishna; Coetzee, William; Goldhamer, David; Hodgson-Zingman, Denice M; Zingman, Leonid V
Sarcolemmal ATP-sensitive potassium (KATP) channels control skeletal muscle energy use through their ability to adjust membrane excitability and related cell functions in accordance with cellular metabolic status. Mice with disrupted skeletal muscle KATP channels exhibit reduced adipocyte size and increased fatty acid release into the circulation. As yet, the molecular mechanisms underlying this link between skeletal muscle KATP channel function and adipose mobilization have not been established. Here, we demonstrate that skeletal muscle-specific disruption of KATP channel function in transgenic (TG) mice promotes production and secretion of musclin. Musclin is a myokine with high homology to atrial natriuretic peptide (ANP) that enhances ANP signaling by competing for elimination. Augmented musclin production in TG mice is driven by a molecular cascade resulting in enhanced acetylation and nuclear exclusion of the transcription factor forkhead box O1 (FOXO1) - an inhibitor of transcription of the musclin encoding gene. Musclin production/secretion in TG is paired with increased mobilization of fatty acids and a clear trend toward increased circulating ANP, an activator of lipolysis. These data establish KATP channel-dependent musclin production as a potential mechanistic link coupling "local" skeletal muscle energy consumption with mobilization of bodily resources from fat. Understanding such mechanisms is an important step toward designing interventions to manage metabolic disorders including those related to excess body fat and associated co-morbidities.
PMCID:4815902
PMID: 26828268
ISSN: 1090-2104
CID: 1932962