Abstract
BackgroundFor the majority of rare clinical missense variants, pathogenicity status cannot currently be classified. Classical homocystinuria, characterized by elevated homocysteine in plasma and urine, is caused by variants in the cystathionine beta-synthase (CBS) gene, most of which are rare. With early detection, existing therapies are highly effective.MethodsDamaging CBS variants can be detected based on their failure to restore growth in yeast cells lacking the yeast ortholog CYS4. This assay has only been applied reactively, after first observing a variant in patients. Using saturation codon-mutagenesis, en masse growth selection, and sequencing, we generated a comprehensive, proactive map of CBS missense variant function.ResultsOur CBS variant effect map far exceeds the performance of computational predictors of disease variants. Map scores correlated strongly with both disease severity (Spearman’s ϱ = 0.9) and human clinical response to vitamin B6 (ϱ = 0.93).ConclusionsWe demonstrate that highly multiplexed cell-based assays can yield proactive maps of variant function and patient response to therapy, even for rare variants not previously seen in the clinic.
Highlights
For the majority of rare clinical missense variants, pathogenicity status cannot currently be classified
To provide a proactive resource to inform the rapid interpretation of genetic variation in cystathionine beta-synthase (CBS), we sought to test all possible missense variants of CBS for functional effects and vitamin B6 remediability
We reimplemented a previously validated humanized yeast model [45,46,47,48, 50], confirming that expression of human CBS from the hORFeome collection restores the ability of a yeast cys4Δ strain to grow without supplementation of glutathione
Summary
For the majority of rare clinical missense variants, pathogenicity status cannot currently be classified. For personalized diagnostic surveillance and therapy, timely and accurate methods to interpret the clinical impact of genetic variants are needed. Over 138,000 exomes have been collected in the Genome Aggregation Database (gnomAD) [1, 2] and 4.6 million coding variants have been discovered. Among these discovered coding variants, 99% are rare, having a minor allele frequency (MAF) below 0.5%. Diverse computational and experimental methods have been developed to predict the functional impact of rare coding variants. At more permissive thresholds that detect 90% of pathogenic variants, fully ~ 30% of pathogenicity predictions were erroneous [6].
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