Genetic and functional analysis of the RYR1 mutation p.Thr84Met revealed a susceptibility to malignant hyperthermia
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The aim of this study was to analyze the genetic and functional role of a novel RYR1 variant c.251 C > T (p.Thr84Met) identified in a patient with muscle weakness demonstrating MH susceptibility.
DNA testing of family members was conducted for assessment of pathogenicity of the genetic variant. For functional analysis, Ca2+ measurement using patient-derived myotubes and p.Thr84Met RYR1-transfected human embryonic kidney (HEK)-293 cells was performed to evaluate reactivity to RYR1 activators. The half-maximal effective concentration (EC50) values of two RYR1 activators, caffeine and 4-chloro-m-cresol (4CmC), were calculated from the acquired dose–response curves. The EC50 was compared between two groups: for myotubes, the control group and the patient, and for HEK-293 cells, WT and p.Thr84Met.
Dose–response curves for caffeine and 4CmC were shifted to the left in both myotubes and HEK-293 cells compared to controls. The 50% effective concentration values for caffeine and 4CmC were significantly lower in both myotubes and HEK-293 cells compared to controls (P < 0.001 for all comparisons).
Our results of functional testing indicated RYR1 hypersensitivity to caffeine and 4CmC. We conclude that the genetic variant was associated with MH susceptibility.
KeywordsMalignant hyperthermia Ryanodine receptor Mutation Calcium release
We thank Professor David H. MacLennan for the generous gift of RYR1/pcDNA.
Compliance with ethical standards
Conflict of interest
This study was supported in part by a Grant-in-Aid (Grant number 16K20098) for Scientific Research from the Japan Society for the Promotion of Science, Japan.
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