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Splicing error in E1alpha pyruvate dehydrogenase mRNA caused by novel intronic mutation responsible for lactic
Manuèle Miné1, Michèle Brivet, Guy Touati
1Laboratoire Centre de Recherche Thérapeutique en Ophtalmologie, Faculté de Médecine Necker, 156 rue de Vaugirard 75015 Paris, France.
Insights
A novel intronic mutation in the E1alpha PDH gene causes aberrant splicing, leading to pyruvate dehydrogenase deficiency. This genetic defect results in developmental delay and lactic acidosis in affected individuals.
Area of Science:
- Genetics
- Molecular Biology
- Biochemistry
Background:
- Pyruvate dehydrogenase (PDH) deficiency is an X-linked metabolic disorder.
- This disorder is characterized by delayed development and lactic acidosis.
- A partial defect in PDH activity underlies the condition.
Observation:
- An intronic point mutation was found in the E1alpha PDH gene of a patient with lactic acidosis.
- Protein analysis showed reduced immunoreactivity of PDH complex subunits.
- Aberrantly spliced mRNA with intron 7 sequences was detected in patient samples.
Findings:
- A G to A substitution in intron 7 (position 26) caused aberrant splicing.
- The mutation created a de novo splicing enhancer motif for SC35.
- This enhancer activated a downstream cryptic 5'-splice site, leading to intron retention.
Implications:
- This intronic mutation is a novel cause of intron retention in human genetic disease.
- Understanding this mechanism can aid in diagnosing and potentially treating PDH deficiency.
- The findings highlight the importance of intronic regions in gene regulation and disease pathogenesis.
Abstract:
An intronic point mutation was identified in the E1alpha PDH gene from a boy with delayed development and lactic acidosis, an X-linked disorder associated with a partial defect in pyruvate dehydrogenase (PDH) activity. Protein analysis demonstrated a corresponding decrease in immunoreactivity of the alpha and beta subunits of the PDH complex. In addition to the normal spliced mRNA product of the E1alpha PDH gene, patient samples contained significant levels of an aberrantly spliced mRNA with the first 45 nucleotides of intron 7 inserted in-frame between exons 7 and 8. The genomic DNA analysis found no mutation in the coding regions but revealed a hemizygous intronic G to A substitution 26 nucleotides downstream from the normal exon 7 5'-splice site. Splicing experiments in COS-7 cells demonstrated that this point mutation at intron 7 position 26 is responsible for the aberrant splicing phenotype, which involves a switch from the use of the normal 5'-splice site (intron 7 position 1) to the cryptic 5'-splice site downstream of the mutation (intron 7 position 45). The intronic mutation is unusual in that it generates a consensus binding motif for the splicing factor, SC35, which normally binds to exonic enhancer elements resulting in increased exon inclusion. Thus, the aberrant splicing phenotype is most likely explained by the generation of a de novo splicing enhancer motif, which activates the downstream cryptic 5'-splice site. The mutation documented here is a novel case of intron retention responsible for a human genetic disease.