RESUMO
We previously reported that microRNA-205-5p (miR-205-5p) is significantly decreased in the ErbB2-overexpressing breast epithelial cell line MCF10A-ErbB2 compared with control cells. In this study, we identified a direct target of miR-205-5p, chloride voltage-gated channel 3 (CLCN3). CLCN3 expression was induced by ErbB2 overexpression; this induced expression was then reduced to control levels by the transfection of the miR-205-5p precursor. In RNA-binding protein immunoprecipitation with Ago1/2/3 antibody, CLCN3 was significantly enriched in 293T embryonic kidney cells with miR-205-5p mimic transfection compared with negative control mimic transfection. In luciferase reporter assays using CLCN3 3'-UTR constructs, the miR-205-5p mimic significantly decreased reporter activity of both wild-type and partial mutant constructs in MCF10A-ErbB2 cells. In contrast, no inhibitory effects of the miR-205-5p mimic were detected using the complete mutant constructs. Since miR-205-5p expression in exosomes derived from MCF10A-neo cells was substantially higher than in exosomes derived from MCF10A-ErbB2 cells, we next investigated whether an exosome-mediated miR-205-5p transfer could control CLCN3 expression. To this end, exosomal miR-205-5p derived from MCF10A-neo cells was functionally transferred to MCF10A-ErbB2 cells, which served to decrease the expression of CLCN3. To assess the roles of CLCN3 in breast cancer, we next performed three-dimensional (3D) spheroid proliferation analyses using MCF10A-ErbB2 cells treated with MCF10A-neo-derived exosomes or CLCN3 shRNA stably expressing SKBR3 and MDA-MB-453 breast cancer cells. Our results showed that both treatment with MCF10A-neo-derived exosome and CLCN3 shRNA expression suppressed 3D spheroid proliferation. Collectively, these novel findings suggest that CLCN3 may be a novel direct target of miR-205-5p and this CLCN3/miR-205-5p interaction may serve a pivotal role in regulating breast cancer cellular proliferation under physiological conditions.
RESUMO
Arsenic (As) and cadmium (Cd) pollution in water is an important global issue. Phytofiltration is an eco-friendly technology that helps clean up pollutants using ornamental plants, such as Micranthemum umbrosum (J.F. Gmel) S.F. Blake. After a seven-day hydroponic experiment, M. umbrosum removed 79.3-89.5% As and 60-73.1% Cd from 0 to 1.0 microg As mL(-1) and 0.3 to 30.0 microg Cd mL(-1) solutions, respectively. For As treatment, root to stem and stem to leaf translocation factors greater than 1.0 indicated that accumulation of As in leaves was large compared to that in stem and roots. However, the accumulation of Cd in roots was higher than that in the leaves and stem. In addition, M. umbrosum completely removed Cd within three days from 0.38 to around 0 microg mL(-1) Cd in the solution when the plant was exchanged daily. Bio-concentration factors (2350 for As and 3027 for Cd) for M. umbrosum were higher than for other As and Cd phytoremediators. The results show that M. umbrosum can be an effective accumulator of Cd and a hyper-accumulator of As, as it can lower As toxicity to a level close to the limit recommended by the World Health Organization (0.01 microg As mL(-1)).
