FIG. 4.

Pdcd10 is a putative target of miR-107 in ^PCMSC. (A) Computational studies showing a putative target site of miR-107 highly conserved in the Pdcd10 mRNA 3′UTR. (B) qRT-PCR showing the successful transfection of MSC for the transgenic expression of miR-107 using the pEZX-miR-107 plasmid. The expression of miR-107 was significantly higher in ¹⁰⁷MSC as compared with the pEZX-miR-Sc plasmid-transfected ^ScMSC as control. The expression of miR-107 in ¹⁰⁷MSC was comparable with ^PCMSC (p>0.05 ¹⁰⁷MSC vs. ^PCMSC). (C) Construction of luciferase construct (D, E). Luciferase activity assay performed by the co-transfection of MSC with pEZX-Luc vector containing (D) Pdcd10 3′UTR or (E) Casp8ap2 3′UTR along with a plasmid encoding miR-107. Significantly decreased luciferase activity was observed on co-transfection with the pEZX-Luc vector containing Pdcd10 3′UTR, whereas no significant decrease in luciferase activity was observed on co-transfection with Casp8ap2 3′UTR. The ratio of luciferase activity was calculated in either the presence or absence of miR-107. (F) Western blot showing significantly higher expression of PDCD10 in ^PCMSC pretreated with anti-miR-107 as compared with ^ScMSC. Fold change was determined in the PDCD10/actin ratio between the Sc siRNA and anti-miR-107-treated cells. (G) Western blot showing significant abrogation of PDCD10 in neonatal cardiomyocytes after the transfection of miR-107 as compared with the non-transfected cardiomyocytes. PDCD10, programmed cell death-10.