1MH Investigation Unit, Clinical Sciences Building, St James University Hospital, Leeds LS9 7TF, UK*Corresponding author
Declaration of interest. This work was supported by donations from the British Malignant Hyperthermia Association.
Accepted for publication: December 4, 2001
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Abstract |
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Methods and results. We screened 200 unrelated MH-susceptible individuals for known CACNA1S mutations. With the aim to characterize further novel mutations at this locus, functionally relevant regions of the gene were also sequenced in 10 unrelated individuals from families where the involvement of other MH susceptibility loci was unlikely. No sequence variations were detected in any of the patients investigated.
Conclusions. Defects in CACNA1S are not a major cause of MH in the UK population. Diagnostic screening of this gene is unlikely to be of value to UK MH patients in the near future.
Br J Anaesth 2002; 88: 5879
Keywords: complications, malignant hyperthermia; genetic factors, hyperthermia
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Introduction |
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Genetic analyses mapped the MH susceptibility trait to the ryanodine receptor locus (RYR1) on chromosome 19q1213.2. This gene encodes the skeletal muscle sarcoplasmic reticulum calcium release channel, a key protein involved in the process of excitationcontraction coupling. To date, over 30 different mutations have been detected in the gene,3 and 15 of these have been functionally characterized using in vitro cellular assays and are considered causative of MH.2 4 It is important to demonstrate that these mutations have a pathogenic effect in a functional assay, as the majority of mutations identified in MH patients are missense mutations (amino acid substitutions), which may have a minimal effect on normal calcium channel function. Approximately 25% of UK MH pedigrees carry one of the 15 causative MH mutations. Whilst other, currently undetected, mutations in RYR1 may be responsible for a large percentage of MH cases (up to 50% show linkage), exclusion of linkage between MH susceptibility and RYR1 has been reported on numerous occasions.5 Linkage studies have implicated other candidate loci on chromosomes 1q, 3q, 5p, 7q and 17q. Therefore MH can be said to be a condition showing considerable allelic and locus heterogeneity.
Monnier and colleagues6 determined linkage to the CACNA1S gene on chromosome 1q with a high degree of probability (two-point LOD score of +4.38) in a large French MH pedigree. CACNA1S encodes the 1 subunit of the dihydropyridine receptor (DHPR), a transverse tubule calcium channel that is tightly coupled to the ryanodine receptor.7 The DHPR functions as the voltage sensor in excitationcontraction coupling. Upon further sequence analysis of MH-susceptible individuals within the pedigree, a G-to-A change at nucleotide 3333 in exon 25 of the CACNA1S gene, substituting arginine for histidine at amino acid 1086, was discovered, which segregated completely with the MH phenotype. This represented the first direct molecular evidence of MH locus heterogeneity. Subsequently Jurkatt-Rott and colleagues8 have found a similar amino acid substitution (Arg1086Cys) in exon 25 that displays partial segregation with the MH phenotype in a German pedigree. Both exons 25 and 26 of CACNA1S encode the intracellular loop that links domains III and IV within the DHPR
1 subunit. Although the functional significance of this region remains unclear, one model suggests that it in some way participates in signalling with the ryanodine receptor.
The aim of this study was to determine whether mutations in the CACNA1S gene predispose the UK population to MH susceptibility. Results of such analyses could have implications for the genetic diagnosis of MH in the future.
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Methods and results |
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A group of MH-susceptible individuals representing 10 UK MH pedigrees was selected. Three exhibited discordance between the RYR1 genotype and MH-susceptibility phenotype, implying the possible presence of a second/alternative causative mutation, or linkage to another susceptibility locus. One represented a pedigree in which a known RYR1 mutation (G7300A) completely segregated with the MH phenotype and therefore served as a CACNA1S mutation negative control. The remaining seven represented pedigrees in which known mutations were excluded and evidence of linkage to chromosome 19 was inconclusive. Genomic DNA samples from all representative MH-susceptible individuals were amplified by the polymerase chain reaction using exon-specific primers for exons 1418 and 2526 (Table 1) and sequenced.
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Comment |
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References |
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2
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