A novel ryanodine receptor mutation and genotype-phenotype correlation in a large malignant hyperthermia New Zealand Maori pedigree

Abstract

Malignant hyperthermia (MH) is a pharmacogenetic disorder that predisposes to a sometimes fatal hypermetabolic reaction to halogenated anaesthetics. MH is considered to originate from abnormal regulation of skeletal muscle Ca(2+) release. Current diagnosis of MH susceptibility (MHS) relies on in vitro contracture testing (IVCT) of skeletal muscle. The ryanodine receptor (RYR1) encoding the major Ca(2+) release channel in the skeletal muscle sarcoplasmic reticulum has been shown to be mutated in a number of MH pedigrees. The large Maori pedigree reported here is the largest MHS pedigree investigated to date and comprises five probands who experienced clinical episodes of MH and 130 members diagnosed by the IVCT. Sequencing of the 15 117 bp RYR1 cDNA in a MHS individual from this pedigree identified a novel C14477T transition that results in a Thr4826 to Ile substitution in the C-terminal region/transmembrane loop of the skeletal muscle ryanodine receptor. This is the first mutation in the RyR1 C-terminal region associated solely with MHS. Although linkage analysis showed strong linkage (max LOD, 11.103 at theta = 0.133) between the mutation and MHS in the pedigree using the standardized European IVCT phenotyping protocol, 22 MHS recombinants were observed. The relationship between the IVCT response and genotype was explored and showed that as IVCT diagnostic cut-off points were made increasingly stringent, the number of MHS discordants decreased with complete concordance between the presence or absence of the C14477T mutation and MHS and MH normal phenotypes, respectively, using a cut-off of 1.2 g tension at 2.0 mM caffeine and 1.8 g tension at 2.0% halothane. Many MHS pedigrees investigated have been excluded from linkage to the RYR1 gene on the basis of a small number of recombinants; however, the linkage analysis reported here suggests that other recombinant families excluded from linkage to the RYR1 gene may actually demonstrate linkage as the number of members tested within the pedigrees increases. The high number of discordants observed using the standardized diagnostic cut-off points is likely to reflect the presence of a second MHS susceptibility locus in the pedigree.

Novel Mutations in C-terminal Channel Region of the Ryanodine Receptor in Malignant Hyperthermia Patients

“…Twenty-three differences in the nucleotide sequence were detected (Table 1). Among these, 19 alterations were silent nucleotide substitutions, 17 of which corresponded to previously reported sequence substitutions (9,15,22,23), and the other three differences were new polymorphisms. None of the 26 previously reported RYR1 mutations associated with MH and/ or CCD (1 -3, 12 -16) was detected in our patients.…”

“…The C-terminal region of RyR1 has been overlooked in the search for MH-associated mutations for some time. Recent sequencing of the complete RyR1 cDNAs from a patient from a Mexican CCD family (14), a patient from a New Zealand Maori MH pedigree (15), and a MH patient from a French family (16) revealed a new region responsible for MH / CCD in the channel region of RyR (MH/ CCD region 3). The novel mutation L4838V in RYR1 found in our MH patients is the first mutation to be located in the transmembrane segment of the channel region, which can be in a broad sense categorized in the MH/CCD region 3.…”

“…Furthermore, deduced amino acid at position 1832 of RyR1 is Glycine in the human genomic DNA sequence database (accession number AC011469 of DDBJ, EMBL and Genbank). The Q3756E mutation was found only in patient #86, and it was also reported to be a polymorphism not associated with MH (15). Therefore, these two alterations, A1832G and Q3756E, are unlikely to be associated with the MH symptoms.…”

The Genetics of Malignant Hyperthermia

“…When more stringent definitions were required to produce an MHS diagnosis from IVCT results, fewer individuals were discordant. When MHS was diagnosed only at an IVCT threshold of 1.2 g contracture in the presence of 2.0 mM caffeine and 1.8 g tension in the presence of 2% halothane, there was no discordance between diagnosis by IVCT and by genotype in this family [19]. Unfortunately, most families do not include such a large number of well-studied individuals.…”

“…In some cases, discordance has been resolved by redefining the cut-off values for the contracture test results [18,19]. This is a reasonable approach if the family is large enough to evaluate linkage with MHS because the definition of positive contracture threshold was based on a large population of subjects.…”