title: Quantitative analysis of CAPN3 transcripts in LGMD2A patients: involvement of nonsense-mediated mRNA decay.
abstract: Limb girdle muscular dystrophy type 2A (LGMD2A) is caused by single or small nucleotide changes widespread along the CAPN3 gene, which encodes the muscle-specific proteolytic enzyme calpain-3. About 356 unique allelic variants of CAPN3 have been identified to date. We performed analysis of the CAPN3 gene in LGMD2A patients at both the mRNA level using reverse transcription-PCR, and at the DNA level using PCR and denaturing high performance liquid chromatography. In four patients, we detected homozygous occurrence of a missense mutation or an in-frame deletion at the mRNA level although the DNA was heterozygous for this mutation in conjunction with a frame-shift mutation. The relationship observed in 12 patients between the quantity of CAPN3 mRNA, determined using real-time PCR, and the genotype leads us to propose that CAPN3 mRNAs which contain frame-shift mutations are degraded by nonsense-mediated mRNA decay. Our results illustrate the importance of DNA analysis for reliable establishment of mutation status, and provide a new insight into the process of mRNA decay in cells of LGMD2A patients.
authors.affiliation: University Hospital Brno, Centre of Molecular Biology and Gene Therapy, Brno, Czech Republic.
authors.forename: Kristýna
authors.initials: K
authors.lastname: Stehlíková
authors.forename: Eva
authors.initials: E
authors.lastname: Zapletalová
authors.forename: Jana
authors.initials: J
authors.lastname: Sedlácková
authors.forename: Markéta
authors.initials: M
authors.lastname: Hermanová
authors.forename: Petr
authors.initials: P
authors.lastname: Vondrácek
authors.forename: Tat'ána
authors.initials: T
authors.lastname: Maríková
authors.forename: Radim
authors.initials: R
authors.lastname: Mazanec
authors.forename: Josef
authors.initials: J
authors.lastname: Zámecník
authors.forename: Stanislav
authors.initials: S
authors.lastname: Vohánka
authors.forename: Jirí
authors.initials: J
authors.lastname: Fajkus
authors.forename: Lenka
authors.initials: L
authors.lastname: Fajkusová
mesh_terms: D000293:Adolescent; D000328:Adult; D015153:Blotting, Western; D002154:Calpain / Q000235:genetics*; D002648:Child; D002675:Child, Preschool; D018389:Codon, Nonsense / Q000235:genetics*; D004247:DNA / Q000096:biosynthesis / Q000235:genetics; D005260:Female; D016368:Frameshift Mutation / Q000235:genetics; D017353:Gene Deletion; D005838:Genotype; D006801:Humans; D007223:Infant; D007438:Introns / Q000235:genetics; D008297:Male; D009124:Muscle Proteins / Q000235:genetics*; D049288:Muscular Dystrophies, Limb-Girdle / Q000235:genetics*; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D018389:Codon, Nonsense; D009124:Muscle Proteins; D012333:RNA, Messenger; D004247:DNA; C105884:CAPN3 protein, human; D002154:Calpain