# MDS/AML-associated DDX41 helicase facilitates homologous recombination repair by potentially resolving R-loops

## TL;DR

This study shows that the DDX41 protein helps repair DNA by resolving R-loops, and its malfunction may lead to blood cancers like MDS and AML.

## Contribution

The study reveals a novel role for DDX41 in resolving R-loops during homologous recombination repair, linking its dysfunction to genomic instability in MDS/AML.

## Key findings

- DDX41 deficiency increases genomic instability and DNA damage in cells.
- DDX41 wild-type resolves R-loops, but the R525H mutant fails to do so.
- DDX41 supports homologous recombination repair by resolving R-loops.

## Abstract

DDX41 mutations are associated with myeloid neoplasms, including myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML), and missense mutant R525H is seen in 67% of patients; however, its molecular pathogenesis is unknown. Using DDX41 knockout (KO) cells, we found that these cells were sensitive to bleomycin, camptothecin, and UV. DDX41 deficiency led to increased genomic instability, indicated by elevated DNA double-strand breaks (DSBs) and comet tails. We found that R-loop formation increased in DDX41–KO cells. DDX41 wild-type (WT) protein resolved DNA:RNA hybrid of R-loops in vitro, but the mutant R525H failed. DDX41–R525H expressing cells were sensitive to DNA damage agents and had significantly more R-loops than DDX41–WT expressing cells. Interestingly, DDX41 colocalized with DSB marker γH2AX and R-loop marker S9.6, and knockdown of DDX41 in the U2OS GFP reporter cells resulted in reduced homologous recombination (HR) repair. Moreover, increased and prolonged RPA and reduced RAD51 foci were found in DDX41 KO and DDX41–R525H expressing cells, indicating a defect in the transition from end resection to RAD51 filament assembly. Overall, our results suggest that DDX41 utilizes its unwinding activity to resolve R-loops, which may play a key role in HR-based repair, and dysregulation of this pathway may lead to MDS/AML.

Graphical Abstract