R-loops Function as a Balance during Somatic Cell Reprogramming

R-loop induces genome instability and regulates gene expression. However, regulatory mechanism of R-loop in stem cell biology remains unclear. Here, we mapped the profiling of R-loops during reprogramming process and revealed that R-loops change prior to gene expression and are mainly accumulated and enriched at terminator of genes. Disrupting homeostasis of R-loops via RNaseH1 loss-of-function blocks reprogramming in a medium independent manner. Interestingly, Sox2 but not other Yamanaka factors overcomes the inhibitory effects of RNaseH1 loss on reprogramming. Sox2 forms a complex with Ddx5 and R-loop in cells and common targets for Sox2 and Ddx5 at R-loops are mainly associated with pluripotent genes. Furthermore, Sox2 inhibits the resolving activity of RNA helicase Ddx5 at R-loops to facilitate reprogramming. Together, these results indicate that the importance of R-loops balance for reprogramming and shed light on the regulation Sox2/Ddx5 on R-loops during reprogramming. Overall design: Gene expression profiling of reprogrammed cells with OSKM at different time points. Gene expression profiling of reprogrammed cells with OSKM plus control/RNaseH1/RNaseH1(D209N) at different time points. R-loops mapping of reprogrammed cells with OSKM at different time points. R-loops mapping of reprogrammed cells with OSKM plus control/RNaseH1/RNaseH1(D209N) at different time points. Chromatin accessibility analysis of reprogrammed cells with OSKM plus control/RNaseH1/RNaseH1(D209N) at different time points. CLIP-seq to identify Ddx5-associated RNAs.