Saturation mutagenesis enables RAD51D clinical variant interpretation and identifies distinct biochemical functions
Abstract
The tumor suppressor RAD51D is essential for homologous recombination (HR). Pathogenic variants in RAD51D are associated with breast and ovarian cancers. However, most clinical missense variants are of uncertain significance. Here, we report 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. We hypothesize that, paradoxically, the primary function of RAD51D is to slow the ATPase activity of BCDX2, thereby allowing sufficient time and space for RAD51 filament assembly. Together, we identify hotspots of deleterious RAD51D variants and uncover the mechanisms by which variants compromise its biochemical functions.
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BibTeX
@article{darrah2026saturation,
title = {{Saturation mutagenesis enables RAD51D clinical variant interpretation and identifies distinct biochemical functions}},
author = {Kristie E. Darrah and Shelby L. Hemker and Yashpal Rawal and Noah J. Goff and Phoebe S. Parker and Gayatri S. Ganesan and Caleb M. Stratton and Katherine G. Oppenheimer and Ella Roberts and Elena Glick and Nicole Banks and Arjun K. Kumar and Silvia Casadei and Matthew W. Snyder and Kara J. Milliron and Sofia D. Merajver and Ana M. Dias and Katherine L. Nathanson and Susan M. Domchek and Lea M. Starita and Shaun K. Olsen and Patrick Sung and Jacob O. Kitzman and Kara A. Bernstein},
journal = {Nature Communications},
year = {2026},
month = oct,
doi = {10.1038/s41467-026-77941-y},
url = {https://www.nature.com/articles/s41467-026-77941-y},
note = {CC BY-NC-ND 4.0},
}
RIS
TY - JOUR
TI - Saturation mutagenesis enables RAD51D clinical variant interpretation and identifies distinct biochemical functions
AU - Kristie E. Darrah
AU - Shelby L. Hemker
AU - Yashpal Rawal
AU - Noah J. Goff
AU - Phoebe S. Parker
AU - Gayatri S. Ganesan
AU - Caleb M. Stratton
AU - Katherine G. Oppenheimer
AU - Ella Roberts
AU - Elena Glick
AU - Nicole Banks
AU - Arjun K. Kumar
AU - Silvia Casadei
AU - Matthew W. Snyder
AU - Kara J. Milliron
AU - Sofia D. Merajver
AU - Ana M. Dias
AU - Katherine L. Nathanson
AU - Susan M. Domchek
AU - Lea M. Starita
AU - Shaun K. Olsen
AU - Patrick Sung
AU - Jacob O. Kitzman
AU - Kara A. Bernstein
PY - 2026
DA - 2026/10/03
JO - Nature Communications
DO - 10.1038/s41467-026-77941-y
UR - https://www.nature.com/articles/s41467-026-77941-y
AB - The tumor suppressor RAD51D is essential for homologous recombination (HR). Pathogenic variants in RAD51D are associated with breast and ovarian cancers. However, most clinical missense variants are of uncertain significance. Here, we report 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. We hypothesize that, paradoxically, the primary function of RAD51D is to slow the ATPase activity of BCDX2, thereby allowing sufficient time and space for RAD51 filament assembly. Together, we identify hotspots of deleterious RAD51D variants and uncover the mechanisms by which variants compromise its biochemical functions.
SN - 2041-1723
ER -