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. 2009 Feb 27;10(4):400–405. doi: 10.1038/embor.2009.9

Figure 2.

Figure 2

The microRNA miR-29a downregulates tristetraprolin. (A) Total RNA was extracted from EpRas and RasXT cells. The levels of TTP as normalized to GAPDH were determined by qPCR. (B) Western blot analysis using whole-cell extracts of EpRas and RasXT cells was performed to monitor the expression of TTP protein. (C) NIH3T3 cells were transfected with miR-scr or miR-29a. Constructs containing the luciferase ORF coupled to the WT and mutated (mut.) versions of the TTP-3′-UTR were used to assess the regulatory impact of miR-29a on TTP. Relative light units (RLUs) are shown. (D) EpRas cells were transfected with miR-scr or miR-29a, and RasXT cells were transfected with LNA-scr or LNA complementary to miR-29a (LNA-29a). Constructs containing the luciferase ORF coupled to the TTP-3′-UTR were used to assess the regulatory impact of endogenous miR-29a on TTP. RLUs are shown. (E) Western blot analysis using whole-cell extracts of EpRas cells and EpRas cells overexpressing miR-29a was performed to monitor the expression of TTP. (F) Western blot analysis using whole-cell extracts of RasXT cells and RasXT cells transfected with LNA-29a was performed to monitor the expression of TTP. LNA, locked nucleic acid; miR-scr, microRNA-scrambled; ORF, open reading frame; qPCR, quantitative PCR; TTP, tristetraprolin; UTR, untranslated region; WT, wild type.