Molecular and functional analysis ofSLC25A20 mutations causing carnitine-acylcarnitine translocase deficiency

Abstract

The enzyme carnitine-acylcarnitine translocase (CACT) is involved in the transport of long-chain fatty acids into mitochondria. CACT deficiency is a life-threatening, recessively inherited disorder of lipid beta-oxidation which manifests in early infancy with hypoketotic hypoglycemia, cardiomyopathy, liver failure, and muscle weakness. We report here the clinical, biochemical, and molecular features of six CACT-deficient patients from Italy, Spain, and North America who exhibited significant clinical heterogeneity. In five patients (Patients 1, 2, 4, 5, and 6) the disease manifested in the neonatal period, while the remaining patient (Patient 3), the younger sibling of an infant who had died with clinical suspicion of fatty acid oxidation defect, has been treated since birth and was clinically asymptomatic at 4.5 years of age. Patients 1 and 4 were deceased within 6 months from the onset of this study, while the remaining four are still alive at 8, 4.5, 3.5, and 2 years, respectively. Sequence analysis of the CACT gene (SLC25A20) disclosed five novel mutations and three previously reported mutations. Three patients were homozygous for the identified mutations. Two of the novel mutations (c.718+1G>C and c.843+4_843+50del) altered the donor splice site of introns 7 and 8, respectively. The 47-nt deletion in intron 8 caused both skipping of exon 8 only and skipping of exons 6-8. Four mutations [[c.159dupT;c.163delA] ([p.Gly54Trp;p.Thr55Ala]) c.397C>T (p.Arg133Trp), c.691G>C (p.Asp231His), and c.842C>T (p.Ala281Val)] resulted in amino acid substitutions affecting evolutionarily conserved regions of the protein. Interestingly, one of these exonic mutations (p.Ala281Val) was associated with a splicing defect also characterized by skipping of exons 6-8. The deleterious effect of the p.Arg133Trp substitution was demonstrated by measuring CACT activity upon expression of the normal and the mutant protein in E. coli and functional reconstitution into liposomes. Combined analysis of clinical, biochemical, and molecular data failed to indicate a correlation between the phenotype and the genotype.

Carnitine-Acylcarnitine Translocase Deficiency: Experience with Four Cases in Spain and Review of the Literature

“…Thirty-one articles describing clinical reports of CACT deficiency were found by searching the PubMed database (Al Aqeel et al 2003;Al-Sannaa and Cheriyan 2010;Brivet et al 1996;Chalmers et al 1997;Costa et al 1999;Costa et al 2003;Fearing et al 2011;Fukushima et al 2013;Galron et al 2004;Geven et al 2007;Huizing et al 1997Huizing et al , 1998Iacobazzi et al 2004a;Iacobazzi et al 2004b;Korman et al 2006;Lee et al 2007;Lopriore et al 2001;Morris et al 1998;Niezen-Koning et al 1995;Nuoffer et al 2000;Ogawa et al 2000;Ogier de Baulny et al 1995;Olpin et al 1997;Pande et al 1993;Parini et al 1999;Pierre et al 2007;Roschinger et al 2000;Rubio-Gozalbo et al 2003;RubioGozalbo et al 2004;Stanley et al 1992;Wang et al 2011;Yang et al 2001). Data from the literature were compiled, compared with the cases reported here, and discussed.…”

“…P2 and P4, two unrelated Spanish cases, both of them with a milder clinical phenotype, presented the same genotype (p.Gly54_Thr55delinsTrpAla/p.Arg178Term). The mutation p.Gly54_Thr55delinsTrpAla has been identified so far in homozygosity in one other Spanish patient with a severe clinical phenotype (Iacobazzi et al 2004a) and in heterozygosis with c.804delG (p.Phe269Serfs*) in one Cape Indian patient with a late onset of symptoms (Wang et al 2011;Fearing et al 2011). The mutation p. Arg178Term has been previously reported in patients of Turkish and North African origin (Rubio-Gozalbo et al 2003;Costa et al 2003;Iacobazzi et al 2004a).…”

“…Despite this, he died of severe gastroenteritis in a Moroccan hospital at 12 months of age. P4 has already been described (Iacobazzi et al 2004a). She is reported here in order to provide updated information regarding her clinical status.…”

“…Now, at 4 years of age, she is asymptomatic with normal psychomotor and growth development (weight 25th percentile, height 90th percentile, BMI 10-25th percentile). Iacobazzi et al (2004a) P3 was the fifth child born to unrelated Moroccan parents after a normal gestation and delivery (BW 3,475 g). Two sisters experienced sudden death at 24 and 48 h of life.…”

“…About 80% of the surviving reported patients were less than 12 years of age. Very few of the surviving patients presented with a milder clinical phenotype (normal physical development with no evidence of cardiac disease or myopathy) and showed homozygosity for private mutations (Morris et al 1998;Iacobazzi et al 2004a;Parini et al 1999;Ijlst et al 2001). Outcomes seem to correlate better with the absence of cardiac disease and a higher long-chain fatty acid oxidation rate in cultured fibroblasts than with residual enzyme activity.…”