Fig 2: MSL1 promotes cell survival in the presence of DNA damage but its upregulation is not dependent on DDR kinases(A) Immunoblots showing increased full-length MSL1 protein expression in HeLa cell nuclear lysate from CPT treatment. UN, untreated. TBP, loading control.(B) The experimental design for the MCA assay. GOI, gene of interest. See STAR Methods for details.(C) MCA assay results from 3 independent experiments. y axis, the relative survival (with DNA damage to without DNA damage) of LN229 cells transfected with different siRNAs. See STAR Methods for the formula for relative survival calculation.(D) HR efficiency in DR-U2OS cells transfected with different siRNAs as measured by flow cytometry from 4 independent experiments.(E) NHEJ efficiency of EJ5-GFP-U2OS cells transfected with different siRNAs as measured by flow cytometry from 3 independent experiments.(F) Immunoblots showing decreased phosphorylation of DDR kinase targets from inhibitor treatments.(G and H) Quantitation of MSL1 mRNA isoforms by qRT-PCR in (G) HeLa cells and (H) U2OS cells with different treatments.

In (F) to (H), the following DDR kinase inhibitors are used: KU55933 (10 µM), KU60019 (10 µM), AZD7648 (3 µM), NU7441 (5 µM), and VE821 (10 µM). The inhibitors were added to media 30 min before addition of CPT. Cells were harvested after 8 h of CPT treatment. In all panels, error bars indicate SEM and statistical significance is determined by one-tailed (C-E) or two-tailed (G and H) t test. NS, not significant; UN, untreated; CPT, 5 µM camptothecin. *p < 0.05; **p < 0.01.