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. 2020 May 11;10(5):58. doi: 10.3390/life10050058

Figure 5.

Figure 5

MiR-145 directly represses DUSP6. Western blot showing reduced DUSP6 levels following miR-145 overexpression in P2 (A) and freshly isolated (B) HACs. (C) Western blot showing increased DUSP6 levels following miR-145 inhibition in freshly isolated HACs. DUSP6 protein bands were quantified by densitometry and are normalized against tubulin and expressed relative to the control (± SEM). * p < 0.05, ** p < 0.01 (A, right hand side panel, B and C, lower panels). In all experiments HACs were transfected with a relevant control (C) or miR-145 precursor or inhibitor, and subsequently cultured in 20% or, where indicated, 1% O2 tension for 44 h. (D) Hela cells were transfected with luciferase reporters containing three perfectly complementary binding sites for miR-145 (3 x miR-145), the DUSP6 3′UTR containing a putative miR-145 binding site (seed matching nucleotides 1018 to 1025; DUSP6 3′UTR), or a mutated seed-matching site (DUSP6 3′UTR miR-145 MUT) downstream the firefly luciferase open reading frame (ORF). Control (‘C’) or miR-145 mimics were cotransfected in the cells. Values were normalized to the levels of renilla luciferase, independently expressed by the same vector and are shown as relative to that obtained for each construct cotransfected with the control miRNA mimic (± SEM). **** p < 0.0001.