Skip to main content
. Author manuscript; available in PMC: 2015 Feb 28.
Published in final edited form as: Nature. 2014 Jun 25;512(7515):431–435. doi: 10.1038/nature13375

Figure 4. Tgif2 is an Essential miR-34a Direct Target and a Pro-Osteoclastogenic Factor.

Figure 4

a, Tgif2 expression was inhibited by pre-miR-34a in osteoclast cultures (n=3). b, Tgif2 expression in WT and 34a-PT-Tg osteoclast cultures (n=3). c, Sequence alignment of the Tgif2 3′UTR. d, A diagram of Tgif2 3′UTR reporters. e, Luciferase readout from WT or mutant Tgif2 3′UTR reporter co-transfected in HEK293 cells with pre-miR-34a or anti-miR-34a (n=3). f-h, Comparison of Tgif2-KO, Tgif2-Het and WT control mice (1.5-month-old, male, n=7). f. Serum CTX-1. g-h, μCT of tibiae. g, Trabecular BV/TV. h, Images of the trabecular bone of the tibial metaphysis (scale bar, 10μm). i, Decreased osteoclast differentiation in Tgif2-KO and Tgif2-Het cultures (n=3). j, Tgif2-KO cultures were resistant to the anti-osteoclastogenic effects of premiR-34a (n=3). i-j, Top, TRAP expression; Bottom, TRAP staining, osteoclast number (black) and resorptive activity (blue). k-l, Tgif2/34a double knockout (DKO) mice were compared with WT, Tgif2-KO or 34a-KO (2-month-old, male, n=4). k, Osteoclast differentiation. l, Serum CTX-1. m, Tgif2 mRNA in RAW264.7 cells following transfection of transcription factors (n=3). n, ChIP of transcription factor binding and H3K4me3 levels at the endogenous Tgif2 promoter in RAW264.7 cells 3d after RANKL treatment (n=6); txn, transcription. o, Transcription factor was co-transfected into 293 cells with its luciferase reporter, together with Tgif2 or a GFP control (n=6). p, Luciferase reporter was transfected into WT, Tgif2-KO or 34a-KO osteoclast cultures (n=6). q-r, NFATc1 mRNA (q, n=3), c-Jun phosphorylation and IκBα degradation (r) in WT, Tgif2-KO or 34a-KO osteoclast cultures. Ratios of p-c-Jun/total-c-Jun and IκBα/β-actin are shown. s, A model for how miR-34a suppresses osteoclastogenesis. Error bars, SD.