BACKGROUND: For 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. METHODS: Damaging 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. RESULTS: Our 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). CONCLUSIONS: We 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.

California Institute for Quantitative Biosciences University of California Berkeley CA 94720 USA

Center for Cancer Systems Biology Dana Farber Cancer Institute Boston MA 02215 USA

Department of Computer Science University of Toronto Toronto ON M5S 3E1 Canada

Department of Genetics Blavatnik Institute Harvard Medical School Boston MA 02115 USA

Department of Medical Biochemistry and Microbiology Uppsala University SE 75123 Uppsala Sweden

Department of Molecular and Cell Biology University of California Berkeley CA 94720 USA

Department of Molecular Genetics University of Toronto Toronto ON M5S 3E1 Canada

Department of Pediatrics and Adolescent Medicine Charles University 1st Faculty of Medicine and General University Hospital Prague 128 08 Praha 2 Czech Republic

Lunenfeld Tanenbaum Research Institute Mount Sinai Hospital Toronto ON M5G 1X5 Canada

The Donnelly Centre University of Toronto Toronto ON M5S 3E1 Canada

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