Experimental validation of the interaction of miR-223, which is expressed highly in naive and memory B cells compared with GC B cells, and targets the transcription factor LMO2. (A) Base-pairing of the 3′UTR LMO2 gene with nucleotides 1-8 of miR-223. This 8-mer is highly conserved across several species and serves as a potential binding site for miR-223. (B) Effects of overexpression of miR-223 in GC lymphoma-derived BJAB cells in 3 separate experiments. The dark gray bars depict expression of LMO2 24 hours after transfection with a scrambled control that does not possess complementarity to the human genome. The light gray bars depict the expression of LMO2 24 hours after transfection with a precursor for miR-223. The expression of LMO2 was consistently lower in the cells treated with the miR-223 precursor (P < .05 in all cases). (C) Relative LMO2 protein expression from a representative experiment (from 3 replicates) transfecting a scrambled control versus a precursor for miR-223 in BJAB cells. (D) Average expression of LMO2 relative to actin over 3 Western blots of BJAB transfected with a scrambled control versus a precursor for miR-223. LMO2 expression is lower in cells treated with miR-223 (P < .05). (E) Luciferase-expressing vectors were coupled to the 3′UTR of the LMO2 gene. The seed sequence mutant construct had consistently diminished miR-223 repression compared with the wild-type construct in 5 separate experiments (P < .05).