Fig. 3.

miR-140-3p targets the CD38 3′-untranslated region (UTR). Luciferase reporter assays were performed in Luc-CD38-3′-UTR constructs containing wild-type or mutated (Mutated) target. A: 2 miR-140-3p targets on CD38 3′-UTR and mutations on each target (top). ORF, open reading frame. Bottom: in human embryonic kidney (HEK)-293 cells cotransfected with wild-type Luc-CD38-3′-UTR and miR-140-3p mimic oligonucleotides, luciferase activity [relative light units (RLU)] was marginally reduced compared with cells cotransfected with scrambled-sequence oligonucleotides; in HEK-293 cells cotransfected with target-mutated Luc-CD38-3′-UTR and miR-140-3p mimic oligonucleotides, inhibition of luciferase activity was partially reversed (n = 3). B: in NIH 3T3 cells cotransfected with wild-type Luc-CD38-3′-UTR and miR-140-3p mimic oligonucleotides, luciferase activity was marginally reduced compared with cells cotransfected with scrambled-sequence oligonucleotides. When the first miR-140-3p target on Luc-CD38-3′-UTR was mutated (mutated 1), inhibition of luciferase activity was completely reversed. When both miR-140-3p targets on Luc-CD38-3′-UTR were mutated (mutated 1+2), luciferase activity was significantly elevated in the presence of miR-140-3p mimic oligonucleotides (n = 4). C: in NAASM and AASM cells transfected with miR-140-3p mimic (mimic 140), CD38 mRNA levels were comparably attenuated at 0 h (NA-0 and A-0; following TNF-α removal and transcriptional arrest) compared with cells transfected with control oligonucleotides. There were no further reductions in CD38 mRNA levels at 6, 12, or 24 h (NA-24 and A-24) following transcriptional arrest in mimic-transfected cells of either group. *P < 0.05 vs. wild-type or mutated vector cotransfected with scramble oligonucleotide. **P < 0.05 vs. wild-type vector cotransfected with miR-140-3p mimic oligonucleotide. ActD, actinomycin D.