Short/branched-chain acyl-CoA dehydrogenase deficiency due to an IVS3+3A>G mutation that causes exon skipping

Abstract

Short/branched-chain acyl-CoA dehydrogenase deficiency (SBCADD) is an autosomal recessive disorder of L: -isoleucine catabolism. Little is known about the clinical presentation associated with this enzyme defect, as it has been reported in only a limited number of patients. Because the presence of C5-carnitine in blood may indicate SBCADD, the disorder may be detected by MS/MS-based routine newborn screening. It is, therefore, important to gain more knowledge about the clinical presentation and the mutational spectrum of SBCADD. In the present study, we have studied two unrelated families with SBCADD, both with seizures and psychomotor delay as the main clinical features. One family illustrates the fact that affected individuals may also remain asymptomatic. In addition, the normal level of newborn blood spot C5-acylcarnitine in one patient underscores the fact that newborn screening by MS/MS currently lacks sensitivity in detecting SBCADD. Until now, seven mutations in the SBCAD gene have been reported, but only three have been tested experimentally. Here, we identify and characterize an IVS3+3A>G mutation (c.303+3A>G) in the SBCAD gene, and provide evidence that this mutation is disease-causing in both families. Using a minigene approach, we show that the IVS3+3A>G mutation causes exon 3 skipping, despite the fact that it does not appear to disrupt the consensus sequence of the 5' splice site. Based on these results and numerous literature examples, we suggest that this type of mutation (IVS+3A>G) induces missplicing only when in the context of non-consensus (weak) 5' splice sites. Statistical analysis of the sequences shows that the wild-type versions of 5' splice sites in which +3A>G mutations cause exon skipping and disease are weaker on average than a random set of 5' splice sites. This finding is relevant to the interpretation of the functional consequences of this type of mutation in other disease genes.

Features of 5′-splice-site efficiency derived from disease-causing mutations and comparative genomics

“…It has been shown that 5Јss with disease-causing +3 A-to-G mutations exhibit two distinct features: (1) They are intrinsically weak (Madsen et al 2006); and (2) they frequently have nonconsensus nucleotides at positions +4 and +5 (for confirmation, see Fig. 4; Ohno et al 1999;Madsen et al 2006). Figure 4 also shows that 5Јss with G at position +3 and SNPs with A/G alleles at position +3 prefer consensus nucleotides at +4 and +5, in agreement with the second observation.…”

“…For this study, we used 595 disease-causing 5Јss mutations from the HGMD database (Stenson et al 2003) plus another independent set (Madsen et al 2006), excluding mutations that affect the nearly invariant +1 and +2 positions.…”

“…Thus, it is not obvious why the A to G mutations at +3 in genes such as ACADSB (also named SBCAD), BRCA1, APC, and NF1 can cause genetic diseases (Madsen et al 2006). It has been shown that 5Јss with disease-causing +3 A-to-G mutations exhibit two distinct features: (1) They are intrinsically weak (Madsen et al 2006); and (2) they frequently have nonconsensus nucleotides at positions +4 and +5 (for confirmation, see Fig. 4; Ohno et al 1999;Madsen et al 2006).…”

New splicing mutations in propionic acidemia

“…1). Madsen et al (2006) recently pointed out that a +3A>G mutation causes exon skipping only in the context of a suboptimal 5¢ splice site, with non-conserved bases mainly at positions +4 and +5, in order to ensure efficient base pairing to U1 snRNA during the splicing process. In agreement with this, the 5¢ splice site of PCCA exon 13 has non-matching nucleotides to U1 snRNA at positions +5 and +6 (Fig.…”

Aspects of Newborn Screening in Isovaleric Acidemia

“…The first patient with SBCADD was reported in 1999 [50]. The ACADSB gene structure was described in 2000 [51], and several mutations in this gene have been reported [49,52,53]. In individuals with this disease, urine analysis reveals marked elevations of 2-methylbutyrylglycine [51,54].…”