FIGURE 6.

Knockdown of ZBTB11 induces R‐loop accumulation and DNA damage to suppress the growth of BC cells in a DDX1‐dependent manner. (A) Rescue of DDX1 expression in ZBTB11 knockdown BC cells was achieved by transfecting a plasmid expressing Flag‐tagged DDX1. The expressions of ZBTB11, Flag‐tagged DDX1, and γH2AX in DDX1‐rescued BC cells were detected by Western blot analysis. (B) R‐loop in ZBTB11 knockdown BC cells with DDX1 rescue was determined by Dot blot using S9.6 antibody. (C) R‐loop and DNA damage in ZBTB11 knockdown BC cells with DDX1 rescue were determined by immunofluorescence, with antibodies against S9.6 and γH2AX, respectively. Scale bar, 10 μm. (D, E) The proliferation and colony formation ability of DDX1‐rescued ZBTB11 knockdown BC cells were evaluated by MTT assay and colony formation assay, respectively. The relative cell growth curves and numbers of colonies are shown. (F) Overexpression of RNase H1 in ZBTB11 knockdown BC cells was achieved by transfecting a plasmid expressing HA‐tagged RNase H1. The expressions of ZBTB11 and HA‐tagged RNase H1 in RNase H1‐overexpressed BC cells were detected by Western blot analysis. (G, H) The proliferation and colony formation ability of RNase H1‐overexpressed ZBTB11 knockdown BC cells were evaluated by MTT assay and colony formation assay, respectively. The relative cell growth curves and numbers of colonies are shown. Statistical significance was assessed using one‐way or two‐way analysis of variance.