Fig. 2.

NRP-2 expression is regulated by miR-331-3p in GBM cells. a Schematic representation of the NRP-2 mRNA, showing the 3′-UTR containing six predicted miR-331-3p binding sites (TargetScan version 6.2, June 2012). Nucleotide positions of miR-331-3p seed sequence sites are shown within the 3′-UTR, with boxes shaded light for non-conserved sites and dark for conserved sites. b Luciferase reporter assay using U-251 MG and U-373 MG cells co-transfected with full length NRP-2 3′-UTR luciferase reporter gene plasmid DNA and either miR-331-3p or miR-NC. For each sample, firefly luciferase activity was normalized to Renilla luciferase activity, and data is expressed relative to each cell line cotransfected with reporter plasmid DNA and miR-NC. Data for each reporter construct is expressed relative to miR-NC transfected cells. Error bars represent standard deviations; *p < 0.05. c Western blotting analysis of NRP-2 protein expression in U-251 MG, U-87 MG and U-373 MG GBM cells 48 h after miR-331-3p or miR-NC transfection. α-tubulin is included as a loading control