title: Mutated p.4894 RyR1 function related to malignant hyperthermia and congenital neuromuscular disease with uniform type 1 fiber (CNMDU1).
abstract: BACKGROUND
  Ryanodine receptor 1 (RyR1) is a Ca(2+) release channel located in the sarcoplasmic reticulum membrane of skeletal muscle. More than 200 variants in RyR1 have been identified in DNA from patients with malignant hyperthermia (MH) and congenital myopathies; only 30 have been sufficiently studied so as to be identified as MH-causative mutations. The Ala4894Thr RyR1 variant was found in a Japanese patient with susceptibility to MH, and the Ala4894Pro variant in a rare case of myopathy: congenital neuromuscular disease with uniform type 1 fiber (CNMDU1). We hypothesized that different Ala4894 variants of RyR1 cause different pathophysiological changes that are identifiable by having differing pharmacological sensitivities to RYR1 agonists.
  
  
  METHODS
  Expression vector with a mutation in RYR1 corresponding to the Ala4894Thr, Ala4894Pro, Ala4894Ser, or Ala 4894Gly variant of human RyR1 was transfected into human embryonic kidney 293 cells. At 72 hours after transfection, we determined the intracellular Ca(2+) changes induced by caffeine and 4-chloro-m-cresol (4CmC), in the presence or absence of dantrolene.
  
  
  RESULTS
  Ala4894Thr-transfected cells and Ala4894Ser-transfected cells were more sensitive to caffeine than the wild type, and Ala4894Thr-transfected cells were also more sensitive to 4CmC than the wild type, whereas Ala4894Pro-transfected cells had no response to caffeine or 4CmC. Ala4894Gly-transfected cells were significantly less sensitive to caffeine than the wild type. In addition, the responses of Ala4894Thr-transfected cells and Ala4894Ser-transfected cells to caffeine were suppressed by dantrolene.
  
  
  CONCLUSIONS
  We concluded that different Ala4894 variants of RyR1 lead to different agonist/antagonist sensitivities, which may predict differing RYR1 functionality during excitation-contraction coupling and sensitivity to MH. The hypersensitive Ala4894Thr-RyR1 is associated with MH and the poorly functional Ala4894Pro-RyR1 with CNMDU1.
authors.affiliation: Department of Anesthesiology and Critical Care, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8551, Japan. tsakharaki@gmail.com
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Haraki
authors.forename: Toshimichi
authors.initials: T
authors.lastname: Yasuda
authors.forename: Keiko
authors.initials: K
authors.lastname: Mukaida
authors.forename: Takako
authors.initials: T
authors.lastname: Migita
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Hamada
authors.forename: Masashi
authors.initials: M
authors.lastname: Kawamoto
mesh_terms: D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D014644:Genetic Variation; D057809:HEK293 Cells; D006801:Humans; D008305:Malignant Hyperthermia / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D018657:Muscle Fibers, Slow-Twitch / Q000187:drug effects / Q000502:physiology*; D016297:Mutagenesis, Site-Directed; D009468:Neuromuscular Diseases / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics* / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: D019837:Ryanodine Receptor Calcium Release Channel; D002110:Caffeine; D002118:Calcium