Extended Data Fig. 3: DYNLL1 chromatin binding suppresses 53BP1 loss-induced restoration of HR in BRCA1 deficient cells. Related to Fig. 3.
From: Dynamics of the DYNLL1–MRE11 complex regulate DNA end resection and recruitment of Shieldin to DSBs
(a) MEFs expressing EGFP-tagged DYNLL1, and EGFP-tagged DYNLL1-FHA domains constructs were exposed to 2 Gy irradiation. 2 h after irradiation cells were fixed and processed for immunofluorescence using a GFP (DYNLL1) antibody. (b, c) MEF p53−/− BRCA1−/− 53BP1−/− cells (B) and COV362 53BP1−/− cells (C) were transfected with EGFP-tagged DYNLL, or EGFP-tagged DYNNL1-FHA constructs. Cells were exposed to 2 Gy irradiation. 2 h later, cells were fixed and processed for immunofluorescence using antibodies against GFP (DYNLL1), RAD51, and γH2AX. (d) COV362 53BP1−/− cells were transfected EGFP-tagged DYNLL, or EGFP-tagged DYNNL1-FHA constructs. Cells were treated with indicated concentrations of Olaparib for 6 days. Percent survival was determined via a cell viability assay. (A-D) n = 3 biologically independent experiments, counting ≥ 100 cells per experiment. Error bars represent the mean±s.e.m. P-values for foci quantification were calculated using two-sided unpaired t-tests. P-value measurements for cell survival curves were assessed by non-regression curve analysis. P values are indicated by nonsignificant (P >0.05), *(P < 0.05), **(P < 0.01), ***(P < 0.001), ****(P<0.0001). Black line in dot plots represent median. Scale bar = 20μm.
