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. 2018 Mar 9;10(4):e8478. doi: 10.15252/emmm.201708478

Figure 1. HDAC3 regulates AKT phosphorylation.

Figure 1

  1. HDACIs inhibited AKT phosphorylation. C4‐2 cells were pre‐treated with 20 μM of CHX for 30 min followed by treatment with pan HDACIs TSA (1 μM), SAHA (5 μM), LBH589 (0.1 μM), or a HDAC6 selective inhibitor Tuba (5 μM) for 24 h prior to Western blot analysis with indicated antibodies. The efficacy of CHX was evident by blockade of induction of FBP1 expression by HDACIs as reported (Yang et al, 2017).
  2. The knockdown efficiency of each member of class I/II HDACs by shRNAs. C4‐2 cells were stably infected with control or gene‐specific shRNAs for 4 days and harvested for RT–qPCR. Expression of each gene was first normalized to the level of GAPDH, and then the expression level of each gene in gene knockdown cells was normalized by that in control knockdown cells. The shRNA knockdown efficiency was determined by subtracting the normalized value from 100%. Data represents means ± SEM. The RT–qPCR was performed in triplicate for each sample.
  3. Control or gene‐specific knockdown C4‐2 cells were serum starved for 24 h and then cultured in regular culture medium for 12 h followed by Western blots for indicated proteins. Western blot bands for total and phosphorylated AKT were quantified and normalized to the quantified values of β‐Tubulin (loading control). The normalized values were further normalized to the value of shC‐infected cells without serum stimulation.
  4. C4‐2 cells were treated with vehicle (DMSO) or HDAC3 inhibitor RGFP966, and at different time points, cells were harvested for Western blots with the indicated antibodies.
  5. The representatives of IHC staining for HDAC3 and AKT S473 phosphorylation in prostate cancer patient specimens; scale bar: 50 μm; scale bar for the inset: 20 μm.
  6. Correlation between expression of AKT S473 phosphorylation and HDAC3 was shown, n = 55, ***P = 1.87e‐07 was performed by Pearson's product‐moment correlation test.

Source data are available online for this figure.