Abstract
Purpose: To investigate the role and mechanism of microRNA-206 (miR-206) in cytoskeleton reorganization in melanoma cells.
 Methods: MiR-206 and RNA helicase p68 (DDX5) expression levels were measured in A375, A875, and HEM-M cells by quantitative real time polymerase chain reaction (qRT-PCR). A DDX5 overexpression cell line was constructed, and DDX5 overexpression, A375, and A875 cells were transfected with miR-206 mimic or DDX5 small interfering RNA (siRNA). Transwell assay was used to assess cell migration and invasion of A375 and A875 cells, while Luciferase reporter assay was used to determine the putative target of miR-206. DDX5, miR-206, vinculin, coronin3, and ezrin expression levels were evaluated by qRT-PCR. Protein expressions of DDX5, vinculin, coronin3, and ezrin were evaluated by western blot analysis.
 Results: DDX5 expression was higher and miR-206 expression lower in A375 and A875 cells when compared to HEM-M cells (p < 0.05). Knockdown of DDX5 and overexpression of miR-206 repressed invasion and migration, and inhibited expression of vinculin, coronin3, and ezrin in A375 and A875 cells (p < 0.05). However, overexpression of DDX5 reversed the effect of miR-206 on cytoskeletal protein expression. Luciferase reporter assay data confirmed that DDX5 is a direct target of miR-206 (p < 0.05).
 Conclusion: MiR-206 suppresses reorganization of the cytoskeleton in melanoma cells by targeting DDX5, and is thus, a promising target for the treatment of melanoma.
Highlights
Malignant melanoma results from proliferation of abnormal melanocytes, has become a common malignant tumor in the past decades, and its incidence and mortality rate are rising [1]
QPCR analysis showed that DDX5 expression was higher in A375 and A875 cells than in HEMM cells (p < 0.01; Figure 1A), whereas miR-206 was lower in A375 and A875 cells than in HEMM cells (p < 0.01; Figure 1B)
These data indicate that DDX5 and miR-206 expression differs in malignant melanoma cells and normal human epidermal melanocytes
Summary
Malignant melanoma results from proliferation of abnormal melanocytes, has become a common malignant tumor in the past decades, and its incidence and mortality rate are rising [1]. It was shown that miRNA affects the cytoskeletal reorganization of cancer cells and regulates the actin cytoskeleton [4]. MiR-206 was found to suppress cell migration and affect the actin cytoskeleton and cell morphology by targeting Coronin 1C in triplenegative breast cancer [5]. MiR-206 inhibited cell invasion and migration of MDA-MB231 breast cancer cells partially through regulation of actin cytoskeleton reorganization, especially filopodia formation [6]. The effect of miR-206 on actin cytoskeleton reorganization, cell morphology, and migration in melanoma cells has not been reported. DDX5 contributed to cytoskeletal reorganization in basal breast cancer cells via modulation of the DDX5-miR-182-actin cytoskeleton pathway [10]. The effects of overexpression and knockdown of miR-206 and DDX5 and their interactions were analyzed in A375 and A875 melanoma cells. Compare multiple groups. p < 0.05 was considered statistically significant
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