Resolving complex splicing outcomes in ALDH3A2 using long-read transcriptome sequencing

Long-read transcriptome sequencing (Iso-Seq) resolves complex aberrant splicing that is difficult to characterize by short-read RNA sequencing. ALDH3A2 encodes fatty aldehyde dehydrogenase, and biallelic pathogenic variants cause Sjögren–Larsson syndrome (SLS), a rare autosomal recessive neurocutaneous disorder. Here, we describe two siblings with typical SLS features (developmental delay, hypotonia, and ichthyosis) together with laryngeal cartilage calcification and recurrent airway obstruction, a previously unreported manifestation. Exome sequencing identified compound heterozygous ALDH3A2 variants: a canonical splice-site variant and a synonymous variant of uncertain DNA-level pathogenicity. Iso-Seq quantified three aberrant isoforms with distinct fates: the exon 5–skipped transcript was a primary target of nonsense-mediated mRNA decay (NMD); the exon 2–skipped transcript carried a premature termination codon but escaped NMD; and the in-frame exon 4–5–skipped transcript escaped NMD and was the only aberrant isoform predicted to yield a translated in-frame protein, despite an internal 109-amino-acid deletion spanning both NAD-binding and catalytic domains. Structural modeling predicted reduced stability and enzymatic function, and ALDH3A2 protein was virtually undetectable in patient cells, supporting pathogenicity. Long-read transcript analysis thus quantitatively prioritizes the dominant protein-producing isoform for functional interrogation, and expands the clinical spectrum of SLS to include laryngeal cartilage calcification.