title: A novel mutation (c.121‑13T>A) in the polypyrimidine tract of the splice acceptor site of intron 2 causes exon 3 skipping in mitochondrial acetoacetyl-CoA thiolase gene.
abstract: Mitochondrial acetoacetyl-CoA thiolase (T2) (gene symbol: ACAT1) deficiency is an autosomal recessive disorder affecting isoleucine catabolism and ketone body utilization. In this study, mutational analysis of an Indian T2-deficient patient revealed a homozygous mutation (c.121‑13T>A) located at the polypyrimidine tract of the splice acceptor site of intron 2, and exon 3 skipping was identified by cDNA analysis using cycloheximide. We made three mutant constructs (c.121‑13T>A, T>C, and T>G substitutions) followed by making a wild-type minigene construct that included an ACAT1 segment from exon 2 to 4 for a splicing experiment. The minigene splicing experiment demonstrated that exon 3 skipping was induced not only by c.121‑13T>A mutation, but also by the other two substitutions. It was difficult to predict the effect of these mutations on splicing using in silico tools, as predictions of different tools were inconsistent with each other. The minigene splicing experiment remains the most reliable method to unravel splicing abnormalities.
authors.affiliation: Department of Pediatrics, Graduate School of Medicine, Gifu University, Gifu 501‑1194, Japan.
authors.forename: Yuka
authors.initials: Y
authors.lastname: Aoyama
authors.affiliation: Department of Pediatrics, Graduate School of Medicine, Gifu University, Gifu 501‑1194, Japan.
authors.forename: Hideo
authors.initials: H
authors.lastname: Sasai
authors.affiliation: Department of Pediatrics, Graduate School of Medicine, Gifu University, Gifu 501‑1194, Japan.
authors.forename: Elsayed
authors.initials: E
authors.lastname: Abdelkreem
authors.affiliation: Department of Pediatrics, Graduate School of Medicine, Gifu University, Gifu 501‑1194, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Otsuka
authors.affiliation: Division of Clinical Genetics, Gifu University Hospital, Gifu 501‑1194, Japan.
authors.forename: Mina
authors.initials: M
authors.lastname: Nakama
authors.affiliation: Department of Pediatrics, Kasturba Medical College, Manipal University, Manipal 576104, India.
authors.forename: Sandeep
authors.initials: S
authors.lastname: Kumar
authors.affiliation: Department of Pediatrics, Kasturba Medical College, Manipal University, Manipal 576104, India.
authors.forename: Shrikiran
authors.initials: S
authors.lastname: Aroor
authors.affiliation: Department of Medical Genetics, Kasturba Medical College, Manipal University, Manipal 576104, India.
authors.forename: Anju
authors.initials: A
authors.lastname: Shukla
authors.affiliation: Department of Pediatrics, Graduate School of Medicine, Gifu University, Gifu 501‑1194, Japan.
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Fukao
mesh_terms: D000101:Acetyl-CoA C-Acetyltransferase / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D017398:Alternative Splicing; D004252:DNA Mutational Analysis; D004789:Enzyme Activation; D005091:Exons; D005260:Female; D050259:Genes, Mitochondrial; D056726:Genetic Association Studies; D020022:Genetic Predisposition to Disease; D006801:Humans; D007223:Infant; D007438:Introns; D009154:Mutation; D022821:RNA Splice Sites
chemical_list: D022821:RNA Splice Sites; D000101:Acetyl-CoA C-Acetyltransferase