Our results reveal a new role of RTEL1 (and Poldip3) in protection against genome instability by preventing excessive R-loop accumulation after replication stress, a condition emerging as a hallmark of cancer and implicated in aging (Halazonetis et al. 2008; Merchut-Maya et al. 2019), thus providing mechanistic insights into the role of RTEL1 in safeguarding proper genome-wide replication and genomic integrity. This evidence concerns the gene RTEL1 and cancer.