MicroRNA-181a Inhibits Activated B-Cell-Like Diffuse Large B-Cell Lymphoma Progression by Repressing CARD11

Figure 3

miR-181a suppresses CARD11 expression in ABC-DLBCLs. (a) QRT-PCR analysis shows relative expression of CARD11 in GCB-like (OCI-Ly19 and SU-DHL-4) and ABC-like (OCI-LY10 and U2932) DLBCL cell lines. Expression was normalized to U6 levels. (b) Representative western blot (left panel) shows CARD11 protein expression in GCB-like (OCI-Ly19 and SU-DHL-4) and ABC-like (OCI-LY10 and U2932) DLBCL cell lines. Right panel shows quantification of CARD11 protein normalized against GAPDH. (c) QRT-PCR analysis shows relative expression of miR-181a (versus GAPDH) in stably transfected miR-181a overexpressing OCI-LY10 and U2932 cells relative to controls transfected with empty vector. (d) Representative western blot shows decreased CARD11 protein expression in miR-181a overexpressing OCI-LY10 and U2932 cell lines than that in controls. (e) Left panel shows schematics of (top) CARD11 mRNA with putative miR-181a binding site in its 3′-untranslated region (3′-UTR) and (b) luciferase reporter constructs with CARD11 mRNA sequences with wild-type or mutated 3′-UTR miR-181a binding sites. Right panel shows relative luciferase activity in OCI-LY10 and U2932 cell lines cells transfected with vector control (blue), CARD11 mRNA with wild-type 3′UTR containing the miR-181a binding site (green) and CARD11 mRNA with 3′UTR containing mutated miR-181a binding site (red). Luciferase activity was measured 48 h after cotransfection. Note: results are expressed as mean ± SEM of three independent experiments; versus vector control group; versus wt-UTR group.