Fig. 6.

ADAR1-dependent regulation of invasion is RNA-editing independent. ADAR1 constructs used for functional assays: OX-P110 (overexpression); ΔCAT-P110; CAT-MUT-P110; and ADAR1 with mutated RNA-binding domains (RBD-mut) and Staufen1 fragments amplified and cloned into pCDNA3 and transfected into 624mel cells. a Their expression, relative to MOCK (Control), was detected by western blot (ADAR1 full length is 110 kDa and lacking the catalytic domain the size is reduced to 64 kDa). β-Actin or vinculin served as controls; b ratio of normalized ADAR1 in each indicated cell line relative to Mock (n-rADAR1 stands for normalized-ratio). The various transfectants were tested for the following: c invasion rate using standardized Boyden chamber assay monitored for 24 h after seeding. Numbers above the bars represent the absolute cell count of invading cells; d ITGB3 expression was examined by qRT-PCR and e by extracellular staining with an anti-ITGB3-FITC-conjugated antibody over isotype control; f pri- and mature miR-22 expression were monitored by qRT-PCR and g, h FoxD1 and Pax6 expression were evaluated using western blot. Results of c, d and f, g represent the mean ± SE of three biologically independent experiments, each performed in triplicates. The numbers in or next to blots indicate molecular marker. Results for a, e, and h are representative of three biologically independent experiments. Asterisks represent P values: *P < 0.05; **P < 0.01 (two-tailed t-test)