Fig. 4

EphA4 is a direct target of miR-106b-5p. a Bioinformatics software (miRNApath, TargetScan, miRDIP, miRanda and miRDB) predict the potential target genes of miR-106b-5p. b Overlapping candidates with genes that are negatively correlated with miR-106b-5p in the TCGA database. c The binding sites of miR-106b-5p on the 3′-UTR of EphA4, and target sequences of EphA4–3′ UTRs were mutated. d Luciferase assay of cells transfected with EphA4–3′UTR-WT or EphA4–3′UTR-MUT reporter together with miR-106b-5p. e Immunoprecipitation of the Ago2/RISC using the Pan-Ago2 antibody in HEMa-LP cells overexpressing miR-106b-5p. IgG was used as a negative control and β-actin was used as an internal control. qRT-qPCR analysis of miR-106b-5p and EphA4 incorporated into RISC in HEMa-LP cells overexpressing miR-106b-5p compared to the levels in the control. U6 and GAPDH was used as an internal control. f RNA levels of EphA4 were detected by qRT-PCR in HEMa-LP cells treated with indicated exosomes or miR-106b-5p mimic. g Western blot analysis identified EphA4 protein expression changes in HEMa-LP cells treated with indicated exosomes or miR-106b-5p mimic. GAPDH was used as a control. h The EphA4 expression profile in human melanoma cell lines (A375, A2058, SK-MEL-28, SK-MEL-1) and human epidermal melanocytes (HEMa-LP). i The correlation of EphA4 mRNA and miR-106b-5p expression in 36 melanoma tissues was negative. j TCGA dataset revealed a significant negative correlation between EphA4 mRNA and miR-106b-5p in melanoma. Data were expressed as the mean ± SD, *P < 0.05, **P < 0.01, ***P < 0.001