Welcome to Gaia! :: View User's Journal | Gaia Journals

Figure optionsDownload full-size imageDownload as PowerPoint slide
MiR-21 directly targets the PDCD4-3′UTR and the 18 bp putative target site could act as the regulatory Darunavir alone
To confirm PDCD4 as a direct target of miR-21 in cervical cancer HeLa cells, we cloned a 478-bp fragment of the PDCD4-3′UTR into a pGL3 vector, downstream of the reporter gene (Fig. 3A). We also constructed a plasmid expressing miR-21 precursor (301 bp) which was amplified from HeLa cells. As shown in Fig. 3B, ectopically expressed miR-21 significantly reduced luciferase activity with wild-type-PDCD4-3′UTR compared to the empty pGL3 vector. This effect could be rescued when we introduced three-point-mutation in the seed sequence (2–4 sites) of pGL3-PDCD4-3′UTR.
Fig. 3.
MiR-21 directly targets the PDCD4-3′UTR. (A) Diagram of PDCD4-3′UTR containing luciferase reporter gene construct and the wt-long-3′UTR fragment we cloned from PDCD4 mRNA in HeLa. The dark region is the putative target site of miR-21. (B) Luciferase Reporter assays. Shown are relative luciferase values normalized to co-transfections with pcDNA6.2-control and pGL3-control. Values are the average ± s.d. of four replicates. ??P nucleotides in the short and long PDCD4-3’UTR fragments are underlined. (D) Luciferase reporter assays as in (B), with Wt-short-3′UTR and Mu-Short-3′UTR. ??P