Novel ALDH18A1 Mutation

Mutations in the gene ALDH18A1 have been identified in patients with neurocutaneous disorders. The gene ALDH18A1 encodes the enzyme P5CS, which facilitates the first step in the de novo synthesis of proline and ornithine. Proline is subsequently used in collagen synthesis (likely the source of cutaneous phenotypes), and ornithine is a key component of the urea cycle (which may explain the neurological deficits in patients).  This project focuses on the functional characterization of a specific ALDH18A1 homozygous variant (T331P) found in four patients within two different, unrelated families, that presented with neurological deficiencies but no cutaneous findings.  We aimed to prove the pathogenicity of this specific variant by studying its impact on the level of collagen and other metabolites, and the localization of the WT and T331P enzyme using ALDH18A1 KO HEK293 cells as a system in the absence of patient-derived fibroblasts.  Our functional studies demonstrated that both WT and T331P P5CS protein are highly stable and expressed at roughly equal levels in the KO cells.   Both WT and T331P enzyme localized to mitochondria, although these organelles appeared enlarged in the T331P P5CS-expressing cells.  Analysis of metabolite levels in WT, ALDH18A1 KO, KO + WT ALDH18A1, and T331P ALDH18A1 was performed using nuclear magnetic resonance (NMR)-based metabolomics.  These experiments showed the reduced ability of the T331P enzyme to increase proline levels, compared to the WT enzyme, when introduced into the KO background.  Collectively, our observations support the likely pathogenic classification of this variant which we expect will be confirmed once experiments on patient-derived tissue can be carried out.  We speculate that the T331P P5CS enzyme provides enough de novo proline synthesis to spare the patients from cutaneous phenotypes, but insufficient ornithine synthesis in neurons to avoid metabolic deficits within the brain.