Detection of a novel ryanodine receptor subtype 1 mutation (R328W) in a malignant hyperthermia family by sequencing of a leukocyte transcript.
Background: To determine whether malignant hyperthermia (MH) susceptibility in a Canadian pedigree is associated with a mutation in the ryanodine receptor subtype 1 (RYR1) gene, the complete RYR1 transcript obtained from the leukocytes of one MH-susceptible family member was sequenced, using a newly...
| Published in: | Anesthesiology Vol. 99; no. 2; pp. 297 - 303 |
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| Main Authors: | , , , , , , |
| Format: | research Journal Article |
| Published: |
Lippincott Williams & Wilkins
Aug2003
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| Online Access: | View this record in EBSCOhost |
| fields | @attributes: recordID: 1 pdfLink: plink: https://search.ebscohost.com/login.aspx?direct=true&db=ccm&AN=137955567&site=ehost-live header: @attributes: shortDbName: ccm uiTerm: 137955567 longDbName: CINAHL Complete uiTag: AN controlInfo: bkinfo: dissinfo: jinfo: jid: 00033022 1V8 jtl: Anesthesiology issn: 00033022 maglogo: N pubinfo: dt: Aug2003 vid: 99 iid: 2 pid: 433 pub: Lippincott Williams & Wilkins place: Baltimore, Maryland artinfo: ui: 137955567 137955567 NLM12883402 137955567 NLM12883402 137955567 ppf: 297 ppct: 6 formats: tig: atl: Detection of a novel ryanodine receptor subtype 1 mutation (R328W) in a malignant hyperthermia family by sequencing of a leukocyte transcript. aug: au: Loke, Julian C P Kraev, Natasha Sharma, Parveen Du, GuoGuang Patel, Leena Kraev, Alexander MacLennan, David H affil: Malignant Hyperthermia Investigation Unit, Banting and Best Department of Medical Research, University of Toronto, Canada sug: subj: Calcium Malignant Hyperthermia Leukocytes Cell Line DNA Amino Acids Pedigree Human RNA Mutation Muscle, Skeletal Drug Effects Caffeine Muscle Contraction Drug Effects Reverse Transcriptase Polymerase Chain Reaction Anesthetics, Inhalation Phosphodiesterase Inhibitors Halothane Validation Studies Comparative Studies Evaluation Research Multicenter Studies ab: Background: To determine whether malignant hyperthermia (MH) susceptibility in a Canadian pedigree is associated with a mutation in the ryanodine receptor subtype 1 (RYR1) gene, the complete RYR1 transcript obtained from the leukocytes of one MH-susceptible family member was sequenced, using a newly developed protocol.Methods: RNA was extracted from leukocytes and converted into complementary DNA. Overlapping fragments of RYR1 complementary DNA were amplified by the polymerase chain reaction and used for double-strand sequencing to find a single mutation likely to be causal of MH susceptibility. Inheritance of the mutation in the family was studied by restriction endonuclease analysis and/or sequencing of genomic DNA and compared to available caffeine halothane contracture test data. The mutation was introduced into rabbit RYR1 complementary DNA, the complementary DNA was expressed in human embryonic kidney line 293 cells, and Ca2+ release by the mutant Ca2+ release channel was measured following the addition of caffeine and halothane.Results: A novel arginine 328 to tryptophan mutation in RYR1 was detected by direct sequencing of the RYR1 transcript from leukocytes of one MH-susceptible individual. A causal role for this mutation in MH is indicated by cosegregation of the mutation with the MH-susceptible phenotype within the family and by the demonstration that the mutant channel has increased sensitivity to both caffeine and halothane.Conclusions: The feasibility of using complete RYR1 transcripts from leukocytes for sequence analysis offers an efficient and noninvasive method for scanning RYR1 for novel mutations. pubtype: Academic Journal doctype: research Journal Article ougenre: Article language: English refInfo: holdings: @attributes: islocal: N |
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