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. 2012 Sep 20;287(46):38505–38514. doi: 10.1074/jbc.M112.400598

FIGURE 5.

FIGURE 5.

3′-UTR luciferase reporter assays. A, 3′-UTR reporter analysis of the vectors containing the PDGFRA or ACVR1 3′-UTR or an MRE containing miR seed sites plus 50–60 nucleotides of flanking sequence, with or without exogenous miR-130b and/or miR-218. The addition of miRs significantly reduced luciferase activity. Luciferase reporter vectors containing the PDGFRA 3′-UTR or microRNA recognition elements show reduced luciferase activity, except for the vector containing the miR-130b MRE. The addition of a morpholino targeting the miR-130b seed site (target protector) restores luciferase activity of the PDGFRA 3′-UTR vector to control levels. B, 3′-UTR reporter analysis of the vectors containing the ACVR2B 3′-UTR or MREs for each of the miR-let-7b or miR-107 binding sites. C, of seven strongly predicted mRNA targets of miR-let-7b, only COL3A failed to show a reduction in luciferase activity. D, effects of 3′-UTRS and MREs on luciferase mRNA levels. Although all of the 3′-UTRs tested reduce luciferase activity and are targeted by miRs (A–C), only 3′-UTR sequences from PDGFRA, and the CPEB1 and CPEB3 MREs, reduced luciferase mRNA levels. The addition of 3′-UTRs for ACVR1, LRIG1, and CPEB4 increased luciferase mRNA levels. Error bars indicate S.D. in A–C and S.E. in D.