Identification of novel genetic interactions with Recql4 in a genome-wide CRISPR/Cas9 synthetic lethal screen.

This study was performed to provide a detailed understanding of the functions and pathways that intersect with the essential DNA replication factor RECQL4. Mutations in RECQL4 cause Rothmund-Thomson Syndrome (RTS), a familial cancer syndrome associated with early onset osteosarcoma in a high proportion of patients. Using murine RECQL4 mutations that closely map to recurrent human RTS mutations, we performed an unbiased genome-wide screen to identify genes that, when deleted, rescued the phenotypes associated with RECQL4 mutation. Overall design: Hoxb8 immortalised R26-CreERT2 Recql4 KO sgKlhdc3 Double Knock-Out (DKO) cells and Recql4 Re-Expressing (RRE) Recql4R26-CreERT2 Recql4 KO sgKlhdc3 cells, stably expressing Cas9 were infected with the BRIE lentiviral Mouse CRISPR Knockout Pooled Library, selected with puromycin for 3 days and grown for 14 days with samples collected at day 0 (is day 3 puromycin) , day 2, day 5 and day 8 to identify guides specifically depleted in the R26-CreERT2 Recql4 KO sgKlhdc3 rescued cells.