miR-155 Inhibits Mouse Osteoblast Differentiation by Suppressing SMAD5 Expression

Figure 3

miR-155 downregulated SMAD5 expression by binding its 3′-UTR site. (a) Plasmid constructs of SMAD5 3′UTR-wt and SMAD5 3′UTR-mut containing either the wild-type or a mutant (marked with red) binding site sequence predicted from a database search; (b) miR-155 and SMAD5 plasmids or scramble control were cotransfected into HEK-293 cells and luciferase activities were determined; (c) SMAD5 protein was detected in MC3T3-E1 cells transfected with either miR-155 or scramble control in control media and p-SMAD5 protein levels were determined in MC3T3-E1 cells transfected with either miR-155 or scramble control after 30 minutes in osteogenic induction media; and (d) expression of SMAD5 mRNA in MC3T3-E1 cells transfected with either miR-155 or scramble control as determined by qRT-PCR; data are presented as mean ± SD; ; versus scramble group.

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