Biology subjects

Plech, M.

Publications and source records attributed to Plech, M..

2 recordsLinked to original sources

bioRxiv2025

Mechanistic Modelling of Recessive Disease through Allelic Integration of Variant Effects

Interpreting the effects of genetic variants remains a major challenge in recessive diseases, where clinical outcomes often depend on interactions between alleles. Multiplex assays of variant effects (MAVEs) measure variant function at scale, but nonlinear relationships with biochemical activity complicate the interpretation of MAVE scores. Here, we describe a broadly applicable approach to estimate enzymatic activities for thousands of genetic variants using a pair of fitness assays conducted at different expression levels, by modelling the nonlinear relationship between activity and fitness. Activity scores from two alleles are then combined into a single pathogenicity metric that captures their joint effect. We applied this approach to adenylosuccinate lyase (ADSL), a purine biosynthesis enzyme mutated in the autosomal recessive disorder ADSL deficiency. Using a yeast-based MAVE, we quantified the functional impact of over 8,000 coding variants. Our framework distinguished pathogenic from benign alleles based on estimated activity, and the integrated pathogenicity score correlated strongly with biochemical measurements from patient-derived cells, outperforming existing computational predictors. This dual innovation--the mechanistic transformation of MAVE data and allelic integration--offers a generalizable strategy for probing enzyme function and interpreting genetic variation in recessive disorders.

genetics↗

bioRxiv2023

Deep mutational scanning quantifies DNA binding and predicts clinical outcomes of PAX6 variants

Nonsense and missense mutations in the transcription factor PAX6 cause a wide range of eye development defects, including aniridia, microphthalmia and coloboma. To understand how changes of PAX6:DNA binding cause these phenotypes, we combined saturation mutagenesis of the paired domain of PAX6 with a yeast one-hybrid (Y1H) assay in which expression of a PAX6-GAL4 fusion gene drives antibiotic resistance. We quantified binding of more than 2,700 single amino-acid variants to two DNA sequence elements. Mutations in DNA-facing residues of the N-terminal subdomain and linker region were particularly detrimental, as were mutations to prolines and to negatively charged residues. Many variants caused molecular gain-of-function effects, including variants in position Ile71 that increased binding to the LE9 enhancer but decreased binding to a SELEX-derived binding site. Benchmarking against known patient variants and applying ACMG/AMP guidelines to variant classification, we obtained supporting to moderate evidence to suggest that 1,306 variants are likely benign, and 977, likely pathogenic. Our analysis shows that most pathogenic mutations in the paired domain of PAX6 can be explained simply by the effects of these mutations on PAX6:DNA association, and establishes Y1H as a generalisable assay for the interpretation of variant effects in transcription factors.

genetics↗