Validation of protein and hybrid interaction. (A) The protein and hybrid interaction is shown by biolayer interferometry. Binding of DPP9 RNA/DNA hybrid (red lines) or dsDNA (blue lines) to high concentration (solid lines) or low concentration (dotted lines) of each protein was measured. These proteins showed more avid binding to RNA/DNA hybrid than to dsDNA. Baseline was recorded from 0 to 1000 sec, association of protein with dsDNA or RNA/DNA hybrid from 1000 to 1600 sec, followed by dissociation. (B) Immunofluorescence staining in primary human fibroblasts showing colocalization of nucleolin and DDX18 (red) with the R-loops stained by S9.6 RNA/DNA hybrid antibody (green). DAPI staining is in blue. (Scale bar) 1 µm.
