title: Functional studies of RYR1 mutations in the skeletal muscle ryanodine receptor using human RYR1 complementary DNA.
abstract: BACKGROUND
  Malignant hyperthermia is associated with mutations within the gene encoding the skeletal muscle ryanodine receptor, the calcium channel that releases Ca from sarcoplasmic reticulum stores triggering muscle contraction, and other metabolic activities. More than 200 variants have been identified in the ryanodine receptor, but only some of these have been shown to functionally affect the calcium channel. To implement genetic testing for malignant hyperthermia, variants must be shown to alter the function of the channel. A number of different ex vivo methods can be used to demonstrate functionality, as long as cells from human patients can be obtained and cultured from at least two unrelated families. Because malignant hyperthermia is an uncommon disorder and many variants seem to be private, including the newly identified H4833Y mutation, these approaches are limited.
  
  
  METHODS
  The authors cloned the human skeletal muscle ryanodine receptor complementary DNA and expressed both normal and mutated forms in HEK-293 cells and carried out functional analysis using ryanodine binding assays in the presence of a specific agonist, 4-chloro-m-cresol, and the antagonist Mg.
  
  
  RESULTS
  Transiently expressed human ryanodine receptor proteins colocalized with an endoplasmic reticulum marker in HEK-293 cells. Ryanodine binding assays confirmed that mutations causing malignant hyperthermia resulted in a hypersensitive channel, while those causing central core disease resulted in a hyposensitive channel.
  
  
  CONCLUSIONS
  The functional assays validate recombinant human skeletal muscle ryanodine receptor for analysis of variants and add an additional mutation (H4833Y) to the repertoire of mutations that can be used for the genetic diagnosis of malignant hyperthermia.
authors.affiliation: Institute of Molecular BioSciences, Massey University, Palmerston North, New Zealand.
authors.forename: Keisaku
authors.initials: K
authors.lastname: Sato
authors.forename: Neil
authors.initials: N
authors.lastname: Pollock
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Stowell
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D002460:Cell Line; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000502:physiology*; D006801:Humans; D008305:Malignant Hyperthermia / Q000209:etiology / Q000235:genetics; D018482:Muscle, Skeletal / Q000502:physiology*; D009154:Mutation / Q000502:physiology*; D011485:Protein Binding / Q000235:genetics; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics* / Q000502:physiology
chemical_list: D018076:DNA, Complementary; D019837:Ryanodine Receptor Calcium Release Channel