Saturation mutagenesis enables RAD51D clinical variant interpretation and identifies distinct biochemical functions

RAD51D

RAD51D loss of function impairing homologous recombination; deleterious variants clustered in the DNA-binding domain and ATPase core

Pathogenic variants in RAD51D, a tumour suppressor gene essential for homologous recombination, are associated with breast and ovarian cancers, yet most clinical missense variants remain of uncertain significance. The authors tested 6,888 RAD51D coding variants for loss of function using a multiplex assay of variant effect. The resulting variant-to-function map perfectly separates known pathogenic and benign variants and is validated by orthogonal homologous recombination and biochemical assays across 70 clinical variants. Variants in the DNA-binding or ATPase core most severely compromise homologous recombination, and the RAD51D-RAD51C interface within the BCDX2 complex proves essential for regulating its ATPase activity. The authors hypothesise that the primary function of RAD51D is to slow BCDX2 ATPase activity, allowing time for RAD51 filament assembly.

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

For a gene whose missense variants are almost all of uncertain significance, a functional map that perfectly separates pathogenic and benign controls provides functional evidence directly usable for reclassification. It will need calibration within the ACMG framework before routine use, and perfect separation on known variants does not guarantee the same performance for rare missense variants outside the critical domains. The resource applies to any RAD51D missense variant detected, whether by exome, genome or panel.

Analysis by Dr Thibaut Benquey