Fig 2: Effect of knockdown of CLDN1 and CLDN11 on the cytotoxicity of anticancer drugs. RERF-LC-AI cells were transfected with siRNAs for a negative control (si-Nega), CLDN1 (si-C1), or CLDN11 (si-C11). (A) The expression levels of CLDN1, CLDN11, and ß-actin in cell lysates were examined by western blotting. The protein levels are shown as a percentage of values in the negative control siRNA (si-Nega). (B) Cytotoxicity was measured using WST-1 assays. (C) The expression levels of ABCB1, ABCC1, ABCG2, ABCC2, and ß-actin in cell lysates were examined by western blotting. A549 cells were used as a positive control in the detection of ABCC2. The protein levels without ABCC2 are shown as a percentage of values in the negative control siRNA (si-Nega). The full-length blot images are shown in Supplementary Fig. S1. n = 3–6. **P < 0.01 and NS, P > 0.05 compared with the negative control siRNA.

Fig 3: Effect of OCLN overexpression on cytotoxicity of anticancer drugs. (A) Mock and OCLN-expressing cells were treated with CDDP or DXR for 48 h. Cell viability was measured by WST-1 assays. (B) The expression of ABCB1, ABCC1, ABCC2, ABCG2, and ß-actin in cell lysates was examined by western blotting. The protein levels of ABC transporters are shown as a percentage of the values in mock cells. The full-length blot images are shown in Supplementary Fig. S4. (C) The mRNA levels of ABCB1, ABCC1, ABCC2, ABCG2 are shown as a percentage of the values in mock cells. n = 3–6. NS, P > 0.05 compared with mock cells.

Fig 4: Increase in the expression of ABCB1 by LM and roscovitine. (A) A549 cells were continuously cultured in the media containing NM or LM. Real-time PCR was performed using primer pairs for ABCB1, ABCC1, ABCC2, ABCG2, and ß-actin. The mRNA levels are represented in percentage to NM. (B) Western blotting was performed using anti-ABCB1, anti-ABCC1, anti-ABCC2, anti-ABCG2, and anti-ß-actin antibodies. The protein levels are represented in percentage to NM. (C) The cells cultured in the NM medium were incubated in the absence and presence of 10 µM roscovitine for 24 h. After Western blotting with anti-ABCB1 and anti-ß-actin antibodies, the protein levels are represented in percentage to the cells without roscovitine. n = 3–4. ** p < 0.01 and * p < 0.05 compared with NM or -roscovitine. NS, p > 0.05.

Fig 5: Decrease in the expression of ABCB1 by SB203580. (A,C) A549 cells were continuously cultured in the media containing NM or LM. The nuclear fraction was applied on SDS-PAGE. Western blotting was performed using anti-Nrf2, anti-HIF-1a, anti-Sp1, anti-c-Jun, anti-p38, and anti-nucleoporin p62 (p62) antibodies. The nuclear protein levels are represented in percentage to NM. (B) A549 cells cultured in the NM medium were incubated in the absence and presence of 10 µM SP600125 (SP) or 400 nM mithramycin (Mith). The protein levels are represented in percentage to the cells without SP or Mith. (D–E) The cells cultured in LM were incubated in the absence and presence of 10 µM SB203580 (SB) for 3 (D), 24 (E), or 6 h (F). (D) Western blotting of the nuclear fraction was performed using anti-p38 and anti-nucleoporin p62 antibodies. (E) Western blotting of the cell lysates was performed using anti-ABCB1 and anti-ß-actin antibodies. (F) Real-time PCR was performed using primer pairs for ABCB1 and ß-actin. The protein and mRNA levels are represented in percentage to the cells without SB. n = 3-4. ** p < 0.01 compared with NM, -SP, -Mith, or -SB. NS, p > 0.05.