Molecular studies of mitochondrial acetoacetyl-coenzyme a thiolase deficiency in the two original families

Abstract

We describe mutations identified in stored skin fibroblast cell lines from two original probands (JB and JM), first reported with 2-methylacetoacetic aciduria, and shown later to have a deficiency of the K(+)-activated enzyme, mitochondrial acetoacetyl-coenzyme A thiolase (T2). JB is homozygous for a 4-base insertion (GCAG) which is derived mutation. The primary mutation is an AG/gt to AG/gc transition at the 5'-splice-junction site in intron 11. An alternative splice site 4 bp downstream (Ggcag/gt) is used which causes a frame shift and replaces 39 C-terminal residues by 70 nonfunctional residues. JM is homozygous for a mutation in the translation-initiation codon (ATG to AAG). By expression analyses the JB mutation (IVS11nt2) causes an unstable T2 polypeptide and the JM mutation (M1K) severely impairs T2 mRNA translation. The JB allele associates with Dutch ancestry (no consanguinity) and the JM allele with Chilean ancestry (distant consanguinity).

Identification of a novel exonic mutation at -13 from 5' splice site causing exon skipping in a girl with mitochondrial acetoacetyl-coenzyme A thiolase deficiency.

“…We have also reported such cases of typical mutations in T2 deficiency (11)(12)(13). We first wondered if the Q272STOP mutation alone would be responsible for the skipping of exon 8.…”

“…The 27-kb human T2 gene, with 12 exons and 11 introns (8), is located on chromosome 1 lq22.3-q23.1 (9). In five different families, we identified seven gene mutations responsible for this disease (10)(11)(12)(13). Among them, four mutations at 5' or 3' splice sites caused skipping ( I I,12) or aberrant splicing (13) of the corresponding exon.…”

“…In five different families, we identified seven gene mutations responsible for this disease (10)(11)(12)(13). Among them, four mutations at 5' or 3' splice sites caused skipping ( I I,12) or aberrant splicing (13) of the corresponding exon. Conservation of nucleotide sequences of 5' and 3' splice sites and the branch site are critical for accurate splicing.…”

Single base substitutions at the initiator codon in the mitochondrial acetoacetyl-CoA thiolase (ACAT1/T2) gene result in production of varying amounts of wild-type T2 polypeptide

“…Although there are examples of initiator non-AUG codons, proteins translated from the non-AUG codons are minor products in comparison with proteins translated from the major AUG codon in all the cases [Florkiewicz and Sommer, 1989;Mehdi et al, 1990;Muralidhar et al, 1994;He et al, 2000]. On the other hand, several initiator codon mutations were reported to be causes of genetic disorders [Pirastu et al, 1984;Moi et al, 1987;Olivieri et al, 1987;Mitchell et al, 1988;John et al, 1989;Patten et al, 1990;Fukao et al, 1993;Breimer et al, 1994;Frank et al, 1999;Umehara et al, 2000;Rendtorff et al, 2001]. We previously identified a c.2T>A mutation in one T2-deficient patient (Patient GK08) [Fukao et al, 1993].…”

“…On the other hand, several initiator codon mutations were reported to be causes of genetic disorders [Pirastu et al, 1984;Moi et al, 1987;Olivieri et al, 1987;Mitchell et al, 1988;John et al, 1989;Patten et al, 1990;Fukao et al, 1993;Breimer et al, 1994;Frank et al, 1999;Umehara et al, 2000;Rendtorff et al, 2001]. We previously identified a c.2T>A mutation in one T2-deficient patient (Patient GK08) [Fukao et al, 1993]. In the present work, we identified another initiator codon mutation (c.2T>C) in a Japanese patient (Patient GK30), and we characterized nine possible single-base substitutions at the initiator codon on the human T2 gene.…”