CDYL1-dependent decrease in lysine crotonylation at DNA double-strand break sites functionally uncouples transcriptional silencing and repair.

Previously, we showed that CDYL1 is recruited to DNA double-strand breaks (DSBs) to promote homologous recombination (HR) repair and foster transcriptional silencing. However, how CDYL1 elicits DSB-induced silencing is not fully understood. Here, we identify a CDYL1-dependent local decrease in the transcriptionally active marks histone lysine crotonylation (Kcr) and crotonylated lysine 9 of H3 (H3K9cr) at AsiSI-induced DSBs, which correlates with transcriptional silencing. Mechanistically, we reveal that CDYL1 crotonyl-CoA hydratase activity counteracts Kcr and H3K9cr at DSB sites, which triggers the eviction of the transcription elongation factor ENL and fosters transcriptional silencing. Furthermore, genetic inhibition of CDYL1 hydratase activity blocks the reduction in H3K9cr and alleviates DSB-induced silencing, whereas HR efficiency unexpectedly remains intact. Therefore, our results functionally uncouple the repair and silencing activity of CDYL1 at DSBs. In a broader context, we address a long-standing question concerning the functional relationship between HR repair and DSB-induced silencing, suggesting that they may occur independently.

Harnessing DNA replication stress to target RBM10 deficiency in lung adenocarcinoma.

The splicing factor RNA-binding motif protein 10 (RBM10) is frequently mutated in lung adenocarcinoma (LUAD) (9-25%). Most RBM10 cancer mutations are loss-of-function, correlating with increased tumorigenesis and limiting the efficacy of current LUAD targeted therapies. Remarkably, therapeutic strategies leveraging RBM10 deficiency remain unexplored. Here, we conduct a CRISPR-Cas9 synthetic lethality (SL) screen and identify ~60 RBM10 SL genes, including WEE1 kinase. WEE1 inhibition sensitizes RBM10-deficient LUAD cells in-vitro and in-vivo. Mechanistically, we identify a splicing-independent role of RBM10 in regulating DNA replication fork progression and replication stress response, which underpins RBM10-WEE1 SL. Additionally, RBM10 interacts with active DNA replication forks, relying on DNA Primase Subunit 1 (PRIM1) that synthesizes Okazaki RNA primers. Functionally, we demonstrate that RBM10 serves as an anchor for recruiting Histone Deacetylase 1 (HDAC1) to facilitate H4K16 deacetylation and R-loop homeostasis to maintain replication fork stability. Collectively, our data reveal a role of RBM10 in fine-tuning DNA replication and provide therapeutic arsenal for targeting RBM10-deficient tumors.