RYR1Sequence Variants in Myopathies: Expression and Functional Studies in Two Families

Abstract

The skeletal muscle ryanodine receptor (RyR1), i.e., the Ca2+ channel of the sarco/endoplasmic reticulum (S/ER), and the voltage-dependent calcium channel Cav1.1 are the principal channels involved in excitation-contraction coupling in skeletal muscle. RYR1 gene variants are linked to distinct skeletal muscle disorders, including malignant hyperthermia susceptibility and central core disease (CCD), mainly with autosomal dominant inheritance, and autosomal recessive myopathies with a broad phenotypic and histopathological spectrum. The age at onset of RYR1-related myopathies varies from infancy to adulthood. We report the identification of four RYR1 variants in two Italian families: one with myopathy and variants c.4003C>T (p.R1335C) and c.7035C>A (p.S2345R), and another with CCD and variants c.9293G>T (p.S3098I) and c.14771_14772insTAGACAGGGTGTTGCTCTGTTGCCCTTCTT (p.F4924_V4925insRQGVALLPFF). We demonstrate that, in patient-specific lymphoblastoid cells, the c.4003C>T (p.R1335C) variant is not expressed and the in-frame 30-nucleotide insertion variant is expressed at a low level. Moreover, Ca2+ release in response to the RyR1 agonist 4-chloro-m-cresol and to thapsigargin showed that the c.7035C>A (p.S2345R) variant causes depletion of S/ER Ca2+ stores and that the compound heterozygosity for variant c.9293G>T (p.S3098I) and the 30-nucleotide insertion increases RyR1-dependent Ca2+ release without affecting ER Ca2+ stores. In conclusion, we detected and functionally characterized disease-causing variants of the RyR1 channel in patient-specific lymphoblastoid cells.

Application Value of Whole Exome Sequencing in Screening and Identifying Pathogenic Genes of Hypopharyngeal Carcinoma

“…In our research, we found that MYH11 has mutations in 7 patients, and we found two pathogenic or possibly pathogenic mutations (rs375159635, rs751495086), which may be related to the pathogenesis of Hypopharyngeal carcinoma. In addition, SDHA is mainly related to gangliomas [28-30], RYR1 is mainly related to myopathy [31,32], and TTN is mainly related to dilated cardiomyopathy [33,34]. Interestingly, we found that these three genes have mutations in more than ve patients, and all of them have two or more pathogenic or possibly pathogenic mutation sites, which indicates their potential in the pathogenesis of Hypopharyngeal carcinoma.…”

Application value of whole exome sequencing in screening and identifying novel mutations of hypopharyngeal cancer

“…In our research, we found that MYH11 mutated in 7 patients, and we found two pathogenic or possibly pathogenic mutations (rs375159635, rs751495086), which may be related to the pathogenesis of hypopharyngeal cancer. In addition, SDHA is mainly related to gangliomas 30 – 32 , RYR1 is mainly related to myopathy 33 , 34 , and TTN is mainly related to dilated cardiomyopathy 35 , 36 . Interestingly, we found that these three genes have mutations in more than five patients, and all of them have two or more pathogenic or possibly pathogenic mutation sites, which indicates their potential in the pathogenesis of hypopharyngeal cancer.…”

Effects of Remimazolam and Propofol on Ca2+ Regulation by Ryanodine Receptor 1 with Malignant Hyperthermia Mutation

“…As previously described, HEK-293 cells transfected with RYR1 /pBI (wild-type) or one of the three RYR1 /pBI mutants (p.Thr84Met [ 14 ], p.Ser2345Arg [ 15 ], or p.Ala4894Thr [ 16 ]) were washed with Hank's balanced salt solution (HBSS) containing 130 mM sodium chloride (NaCl), 5.4 mM potassium chloride (KCl), 20 mM HEPES, 2.5 mM calcium chloride (CaCl 2 ), 1 mM magnesium chloride (MgCl 2 ), and 5.5 mM glucose (pH 7.4). The cells were loaded with 5.0 μ M Fura-2 AM (Dojindo Molecular Technologies, Tokyo, Japan) in HBSS for 1 hour at 37°C and then washed with HBSS.…”