Molecular characterization of the calcium release channel deficiency syndrome

David J. Tester; CS John Kim; Samantha K. Hamrick; Dan Ye; Bailey J. O'Hare; Hannah M. Bombei; Kristi K. Fitzgerald; Carla M. Haglund-Turnquist; Dianne L. Atkins; Luis A. Ochoa Nunez; Ian H. Law; Joel D. Temple; Michael J. Ackerman

doi:10.1172/jci.insight.135952

Molecular characterization of the calcium release channel deficiency syndrome

David J. Tester,¹ CS John Kim,¹ Samantha K. Hamrick,¹ Dan Ye,² Bailey J. O'Hare,¹ Hannah M. Bombei,³ Kristi K. Fitzgerald,⁴ Carla M. Haglund-Turnquist,¹ Dianne L. Atkins,⁵ Luis A. Ochoa Nunez,³ Ian H. Law,³ Joel D. Temple,⁴ and Michael J. Ackerman⁶

Published July 14, 2020 - More info

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Abstract

We identified a novel homozygous duplication involving the promoter region and exons 1-4 of RYR2 that is responsible for highly penetrant, exertion-related sudden deaths/cardiac arrests in the Amish community without an overt phenotype to suggest RYR2-mediated catecholaminergic polymorphic ventricular tachycardia (CPVT). Homozygous RYR2-duplication (RYR2-DUP) induced pluripotent stem cell-cardiomyocytes (iPSC-CMs) were generated from two unrelated patients. There was no difference in baseline Ca²⁺ handling measurements between WT^- and the RYR2-DUP-iPSC-CMs lines. However, compared to WT-iPSC-CMs, both patient lines demonstrated a dramatic reduction in caffeine and isoproterenol (ISO) stimulated Ca²⁺ transient amplitude, suggesting RyR2 loss-of-function. There was a >50% reduction in RYR2 transcript/RyR2 protein expression in both patient iPSC-CMs compared to WT. Delayed afterdepolarization was observed in the RYR2-DUP-iPSC-CMs but not in the WT-iPSC-CMs. Compared to WT-iPSC-CMs, there was a significantly elevated arrhythmic activity in the RYR2-DUP-iPSC-CMs in response to ISO. Nadolol, propranolol, and flecainide reduced erratic activity by 8.5 fold, 6.8 fold, and 2.4 fold, respectively from ISO challenge. Unlike the gain-of-function mechanism observed in RYR2-mediated CPVT, the homozygous multi-exon duplication precipitates a dramatic reduction in RYR2 transcription and RyR2 protein translation, a loss-of-function in calcium handling, and a calcium-induced calcium release apparatus that is insensitive to catecholamines and caffeine.

Molecular characterization of the calcium release channel deficiency syndrome

David J. Tester,¹ CS John Kim,¹ Samantha K. Hamrick,¹ Dan Ye,² Bailey J. O'Hare,¹ Hannah M. Bombei,³ Kristi K. Fitzgerald,⁴ Carla M. Haglund-Turnquist,¹ Dianne L. Atkins,⁵ Luis A. Ochoa Nunez,³ Ian H. Law,³ Joel D. Temple,⁴ and Michael J. Ackerman⁶

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Molecular characterization of the calcium release channel deficiency syndrome

David J. Tester,1 CS John Kim,1 Samantha K. Hamrick,1 Dan Ye,2 Bailey J. O'Hare,1 Hannah M. Bombei,3 Kristi K. Fitzgerald,4 Carla M. Haglund-Turnquist,1 Dianne L. Atkins,5 Luis A. Ochoa Nunez,3 Ian H. Law,3 Joel D. Temple,4 and Michael J. Ackerman6

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David J. Tester,¹ CS John Kim,¹ Samantha K. Hamrick,¹ Dan Ye,² Bailey J. O'Hare,¹ Hannah M. Bombei,³ Kristi K. Fitzgerald,⁴ Carla M. Haglund-Turnquist,¹ Dianne L. Atkins,⁵ Luis A. Ochoa Nunez,³ Ian H. Law,³ Joel D. Temple,⁴ and Michael J. Ackerman⁶