Research Paper Volume 12, Issue 14 pp 14754—14774

Figure 5. DSCAM-AS1 regulated QPRT expression by sponging miRNAs. (A) miRNAs targeting DSCAM-AS1, DCTPP1 and QPRT were predicated using PITA, RNA22, miRmap, mircoT, miRanda, PicTar, and Targetscan softwares. Venn diagram was used to find miRNAs targeting both DSCAM-AS1 and DCTPP1 or QPRT. (B) miRNA-494-3p was predicted to tightly bind to DSCAM-AS1; however, the connection between miRNA-494-3p and the DCTPP1 mRNA seem to be weak. miRNA-150-5p and miRNA-2467-3p were predicted to tightly bind to both DSCAM-AS1 and QPRT mRNA. (C) PCR was performed to detect miRNA-494-3p, miRNA-150-5p and miRNA-2467-3p expression after transfection with their inhibitors and mimics. (D) PCR was performed to detect DCTPP1 or QPRT expression after transfection with miRNA-494-3p, miRNA-150-5p and miRNA-2467-3p inhibitors and mimics. (E) As per the RNA pull-down assay, both miRNA-150-5p and miRNA-2467-3p WT pulled down DSCAM-AS1 from the cell lysate (P<0.001, E), but their MT failed to do so. (F) The association between miRNA-150-5p and miRNA-2467-3p and the QPRT mRNA 3′ UTR was determined using a luciferase reporter assay. Transfection with miRNA-150-5p and miRNA-2467-3p mimics decreased the WT luciferase reporter activity; however, the transfection did not decrease the MT luciferase reporter activity. *P<0.05, **P<0.01 and ***P<0.001 vs. control group.