Loss of BRCA1 and SETX increases cellular dsRNA. (A) Representative confocal images showing HeLa cells transfected with ATTO-594 (red)-labeled dsRNA or RNA–DNA hybrids and stained with the J2 antibody (green). Inset images are magnified to show overlap of J2 signal and ATTO-594 foci. (B) Representative confocal images of HeLa cells transfected with control or BRCA1-targeting siRNAs. After fixation, coverslips were either mock treated or treated with RNase III. J2 signal is shown in red, DAPI in blue. (C) Quantification of mean nuclear J2 intensities shown in B. (D) Same as in B but following SETX depletion. (E) Quantification of mean nuclear J2 intensities shown in D. All box plots show median (box central line), 25% and 75% percentiles (box edges), and 10% and 90% percentiles (whiskers). Data are combined from three biological replicates (n = 3), with at least 75 nuclei scored per condition per experiment. ***, P ≤ 0.001 (by Mann-Whitney U test); ns, P > 0.05. Scale bar is 10 microns; 5 microns for inset images.