High-throughput mapping of 6,888 RAD51D variants identifies distinct biochemical functions needed for homologous recombination and olaparib response

Pathogenic variants in RAD51D are associated with breast and ovarian cancers. However, most clinical missense variants are of unknown significance.

We performed a multiplex assay of variant effect to test 6,888 RAD51D coding variants for loss-of-function. The resulting variant-to-function map perfectly separates known pathogenic and benign variants and is validated by orthogonal HR and biochemical assays across 70 clinical variants. Our screen shows that variants in the DNA- binding or ATPase core most severely compromise HR, and we identify the RAD51D-RAD51C interface within the BCDX2 complex as essential for regulating its ATPase activity.