👤 Kazue Edaki

Transcriptional factors, such as Snail, Slug, and Smuc, that cause epithelial-mesenchymal transition are thought to regulate the expression of Ezrin, Radixin, and Moesin (ERM proteins), which serve as anchors for efflux transporters on the plasma membrane surface. Our previous results using lung cancer clinical samples indicated a correlation between Slug and efflux transporter MRP2. In the current study, we aimed to evaluate the relationships between MRP2, ERM proteins, and Slug in lung cancer cells. HCC827 cells were transfected by Mock and Slug plasmid. Both mRNA expression levels and protein expression levels were measured. Then, the activity of MRP2 was evaluated using CDCF and SN-38 (MRP2 substrates). HCC827 cells transfected with the Slug plasmid showed significantly higher mRNA expression levels of MRP2 than the Mock-transfected cells. However, the mRNA expression levels of ERM proteins did not show a significant difference between Slug-transfected cells and Mock-transfected cells. Protein expression of MRP2 was increased in Slug-transfected cells. The uptake of both CDCF and SN-38 was significantly decreased after transfection with Slug. This change was abrogated by treatment with MK571, an MRP2 inhibitor. The viability of Slug-transfected cells, compared to Mock cells, significantly increased after incubation with SN-38. Thus, Slug may increase the mRNA and protein expression of MRP2 without regulation by ERM proteins in HCC827 cells, thereby enhancing MRP2 activity. Inhibition of Slug may reduce the efficacy of multidrug resistance in lung cancer. Show less

Epithelial-mesenchymal transition (EMT) plays a role in cancer metastasis as well as in drug resistance through various mechanisms, including increased drug efflux mediated by P-glycoprotein (P-gp). In this study, we investigated the activation mechanism of P-gp, including its regulatory factors, during EMT in hepatoblastoma-derived HepG2 cells. HepG2 cells were transfected with SNAI1 using human adenovirus serotype 5 vector. We quantified mRNA and protein expression levels using qRT-PCR and western blot analysis, respectively. P-gp activity was evaluated by uptake assay, and cell viability was assessed by an MTT assay. P-gp protein expression on plasma membrane was higher in SNAI1-transfected cells than in Mock cells, although there was no difference in P-gp protein level in whole cells. Among the scaffold proteins such as ezrin, radixin and moesin (ERM), only radixin was increased in SNAI1-transfected cells. Uptake of both Rho123 and paclitaxel was decreased in SNAI1-transfected cells, and this decrease was blocked by verapamil, a P-gp inhibitor. The reduced susceptibility of SNAI1-transfected cells to paclitaxel was reversed by elacridar, another P-gp inhibitor. Increased expression of radixin during SNAI1-induced EMT leads to increased P-gp membrane expression in HepG2 cells, enhancing P-gp function and thereby increasing drug resistance. Show less