Functional profiling of STXBP1 missense variants using a novel dual-readout fluorometric assay.

STXBP1

STXBP1 missense variants reducing protein stability and binding to syntaxin-1A (STX1A), sometimes with STX1A conformation-specific deficits

STXBP1-related disorders are among the most common genetic neurodevelopmental disorders, and many missense variants remain uncharacterized, with ACMG/AMP criteria having limited discriminatory power for lack of functional data. The authors developed a split-fluorescence-complementation assay that simultaneously measures STXBP1 protein stability and binding to syntaxin-1A, including its closed and open conformations. Across 15 variants (5 benign, 6 pathogenic, 4 associated with an atypically mild presentation), benign variants showed no marked effect, pathogenic ones reduced stability and binding, and mild-phenotype variants displayed intermediate profiles. An exploratory support vector machine classifier correctly classified benign and pathogenic variants in leave-one-out cross-validation, except for one variant, and its score correlated with clinical severity.

Synthesis written by Geno'X. For the full original abstract, please refer to the source publication.

A quantitative functional assay on a gene with a high volume of missense variants provides a directly usable piece of evidence for variants of uncertain significance, provided it is calibrated against controls. Here, 15 variants and an exploratory classifier are not yet enough to set classification thresholds, and the misclassified variant in a mild-phenotype patient illustrates the limit. It is a tool for the interpretation step of exome/genome-derived variants, not a test to deploy in routine.

Analysis by Dr Thibaut Benquey