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. 2012 Apr 23;120(2):249–258. doi: 10.1182/blood-2012-02-408492

Figure 5.

Figure 5

Validation of FBXL20 and USP40 as direct targets of miR-3151. Expression levels of FBXL20 and USP40 as determined by RT-PCR in KG1 cells infected in triplicate with miR-3151 compared relative to scramble control are shown. Forced miR-3151 expression resulted in an 85% decrease of FBXL20 (± SD; A) and a 66% decrease of USP40 expression levels (± SD; B, both P < .001; expression levels are displayed relative to scramble control). (C) Validation of increased miR-3151 expression levels after lentiviral infection. (D) Western blot of FBXL20 expression comparing miR-3151 infected KG1 cells versus scramble control. miR-3151 expression results in elimination of the FBXL20 protein. The effect of miR-3151 on luciferase activity is shown for the FBXL20 (E) and USP40 (F) 3′-UTRs (cloned 3′ of the luciferase gene). The addition of miR-3151 resulted in a decrease of luciferase activity compared with scramble control of 54% for FBXL20 and of 33% for USP40 (± SD; both P < .001). This effect was abrogated after mutation of the respective binding sequences of the predicted miR-3151–binding sites.