Metastable Atrial State Underlies the Primary Genetic Substrate for MYL4 Mutation-Associated Atrial Fibrillation
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Metastable Atrial State Underlies the Primary Genetic Substrate for MYL4 Mutation-Associated Atrial Fibrillation. / Ghazizadeh, Zaniar; Kiviniemi, Tuomas; Olafsson, Sigurast; Plotnick, David; Beerens, Manu E; Zhang, Kun; Gillon, Leah; Steinbaugh, Michael J; Barrera, Victor; Sui, Shannan Ho; Werdich, Andreas A; Kapur, Sunil; Eranti, Antti; Gunn, Jarmo; Jalkanen, Juho; Airaksinen, Juhani; Kleber, Andre G; Hollmén, Maija; MacRae, Calum A.
In: CIRCULATION, Vol. 141, No. 4, 28.01.2020, p. 301-312.Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review
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TY - JOUR
T1 - Metastable Atrial State Underlies the Primary Genetic Substrate for MYL4 Mutation-Associated Atrial Fibrillation
AU - Ghazizadeh, Zaniar
AU - Kiviniemi, Tuomas
AU - Olafsson, Sigurast
AU - Plotnick, David
AU - Beerens, Manu E
AU - Zhang, Kun
AU - Gillon, Leah
AU - Steinbaugh, Michael J
AU - Barrera, Victor
AU - Sui, Shannan Ho
AU - Werdich, Andreas A
AU - Kapur, Sunil
AU - Eranti, Antti
AU - Gunn, Jarmo
AU - Jalkanen, Juho
AU - Airaksinen, Juhani
AU - Kleber, Andre G
AU - Hollmén, Maija
AU - MacRae, Calum A
PY - 2020/1/28
Y1 - 2020/1/28
N2 - BACKGROUND: Atrial fibrillation (AF) is the most common clinical arrhythmia and is associated with heart failure, stroke, and increased mortality. The myocardial substrate for AF is poorly understood because of limited access to primary human tissue and mechanistic questions around existing in vitro or in vivo models.METHODS: Using an MYH6:mCherry knock-in reporter line, we developed a protocol to generate and highly purify human pluripotent stem cell-derived cardiomyocytes displaying physiological and molecular characteristics of atrial cells. We modeled human MYL4 mutants, one of the few definitive genetic causes of AF. To explore non-cell-autonomous components of AF substrate, we also created a zebrafish Myl4 knockout model, which exhibited molecular, cellular, and physiologic abnormalities that parallel those in humans bearing the cognate mutations.RESULTS: There was evidence of increased retinoic acid signaling in both human embryonic stem cells and zebrafish mutant models, as well as abnormal expression and localization of cytoskeletal proteins, and loss of intracellular nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide + hydrogen. To identify potentially druggable proximate mechanisms, we performed a chemical suppressor screen integrating multiple human cellular and zebrafish in vivo endpoints. This screen identified Cx43 (connexin 43) hemichannel blockade as a robust suppressor of the abnormal phenotypes in both models of MYL4 (myosin light chain 4)-related atrial cardiomyopathy. Immunofluorescence and coimmunoprecipitation studies revealed an interaction between MYL4 and Cx43 with altered localization of Cx43 hemichannels to the lateral membrane in MYL4 mutants, as well as in atrial biopsies from unselected forms of human AF. The membrane fraction from MYL4-/- human embryonic stem cell derived atrial cells demonstrated increased phospho-Cx43, which was further accentuated by retinoic acid treatment and by the presence of risk alleles at the Pitx2 locus. PKC (protein kinase C) was induced by retinoic acid, and PKC inhibition also rescued the abnormal phenotypes in the atrial cardiomyopathy models.CONCLUSIONS: These data establish a mechanistic link between the transcriptional, metabolic and electrical pathways previously implicated in AF substrate and suggest novel avenues for the prevention or therapy of this common arrhythmia.
AB - BACKGROUND: Atrial fibrillation (AF) is the most common clinical arrhythmia and is associated with heart failure, stroke, and increased mortality. The myocardial substrate for AF is poorly understood because of limited access to primary human tissue and mechanistic questions around existing in vitro or in vivo models.METHODS: Using an MYH6:mCherry knock-in reporter line, we developed a protocol to generate and highly purify human pluripotent stem cell-derived cardiomyocytes displaying physiological and molecular characteristics of atrial cells. We modeled human MYL4 mutants, one of the few definitive genetic causes of AF. To explore non-cell-autonomous components of AF substrate, we also created a zebrafish Myl4 knockout model, which exhibited molecular, cellular, and physiologic abnormalities that parallel those in humans bearing the cognate mutations.RESULTS: There was evidence of increased retinoic acid signaling in both human embryonic stem cells and zebrafish mutant models, as well as abnormal expression and localization of cytoskeletal proteins, and loss of intracellular nicotinamide adenine dinucleotide and nicotinamide adenine dinucleotide + hydrogen. To identify potentially druggable proximate mechanisms, we performed a chemical suppressor screen integrating multiple human cellular and zebrafish in vivo endpoints. This screen identified Cx43 (connexin 43) hemichannel blockade as a robust suppressor of the abnormal phenotypes in both models of MYL4 (myosin light chain 4)-related atrial cardiomyopathy. Immunofluorescence and coimmunoprecipitation studies revealed an interaction between MYL4 and Cx43 with altered localization of Cx43 hemichannels to the lateral membrane in MYL4 mutants, as well as in atrial biopsies from unselected forms of human AF. The membrane fraction from MYL4-/- human embryonic stem cell derived atrial cells demonstrated increased phospho-Cx43, which was further accentuated by retinoic acid treatment and by the presence of risk alleles at the Pitx2 locus. PKC (protein kinase C) was induced by retinoic acid, and PKC inhibition also rescued the abnormal phenotypes in the atrial cardiomyopathy models.CONCLUSIONS: These data establish a mechanistic link between the transcriptional, metabolic and electrical pathways previously implicated in AF substrate and suggest novel avenues for the prevention or therapy of this common arrhythmia.
KW - Animals
KW - Animals, Genetically Modified/genetics
KW - Atrial Fibrillation/genetics
KW - Cell Line
KW - Connexin 43/genetics
KW - Gene Knockout Techniques
KW - Heart Atria/metabolism
KW - Human Embryonic Stem Cells/metabolism
KW - Humans
KW - Induced Pluripotent Stem Cells/metabolism
KW - Mutation
KW - Myocytes, Cardiac/metabolism
KW - Myosin Light Chains/genetics
KW - Zebrafish
KW - Zebrafish Proteins/genetics
U2 - 10.1161/CIRCULATIONAHA.119.044268
DO - 10.1161/CIRCULATIONAHA.119.044268
M3 - SCORING: Journal article
C2 - 31735076
VL - 141
SP - 301
EP - 312
JO - CIRCULATION
JF - CIRCULATION
SN - 0009-7322
IS - 4
ER -