DEAD box helicase 41 (DDX41) mutations are the most prevalent predisposition to familial myelodysplastic syndrome (MDS). However, the precise roles of these variants in the pathogenesis of MDS have yet to be elucidated. Here, we discovered a novel mechanism by which DDX41 contributes to R-loop-induced DNA damage responses (DDR) in cooperation with the m6A-METTL complex (MAC) and YTHDC1 using DDX41 knockout (KO) and DDX41 knock-in (KI, R525H, Y259C) cell lines as well as primary samples from MDS patients. Compared to wild type (WT), DDX41 KO and KI led to increased levels of m6A RNA methylated R-loop. Interestingly, we found that DDX41 regulates m6A/R-loop levels by interacting with MAC components. Further, DDX41 promoted the recruitment of YTHDC1 to R-loops by promoting the binding between METTL3 and YTHDC1, which was dysregulated in DDX41-deficient cells, contributing to genomic instability. Collectively, we demonstrated that DDX41 plays a key role in the physiological control of R-loops in cooperation with MAC and YTHDC1. These findings provide novel insights into how defects in DDX41 influence MDS pathogenesis and suggest potential therapeutic targets for the treatment of MDS.

DEAD 框解旋酶 41 (DDX41) 突变是家族性骨髓增生异常综合征 (MDS) 最常见的易感因素。然而，这些变异在MDS发病机制中的确切作用尚未阐明。在这里，我们发现了一种新机制，通过该机制，DDX41 与 m6A-METTL 复合物 (MAC) 和 YTHDC1 配合使用DDX41敲除 (KO) 和DDX41敲入 (KI， R525H、Y259C）细胞系以及来自 MDS 患者的原始样本。与野生型 (WT) 相比，DDX41 KO 和 KI 导致 m6A RNA 甲基化 R 环水平增加。有趣的是，我们发现 DDX41 通过与 MAC 成分相互作用来调节 m6A/R 环水平。此外，DDX41 通过促进 METTL3 和 YTHDC1 之间的结合，促进 YTHDC1 募集到 R 环，而 YTHDC1 在DDX41缺陷细胞中失调，导致基因组不稳定。总的来说，我们证明了 DDX41 与 MAC 和 YTHDC1 合作在 R 环的生理控制中发挥着关键作用。这些发现为 DDX41 缺陷如何影响 MDS 发病机制提供了新的见解，并提出了治疗 MDS 的潜在治疗靶点。