Fig. 6: HDAC8 controls HIF1α levels and JNK activation through TRAF7.
From: Hypoxia-induced conversion of sensory Schwann cells into repair cells is regulated by HDAC8
a, b HIF1α (a) or c-Jun (b) Western blot in lysates of rat SCs cultured in conditions mimicking the conversion into the repair phenotype in normoxia (a) or hypoxia (b) and incubated with lentiviruses carrying a TRAF7 shRNA (T7sh) or control shRNA (Csh), and quantification normalized to GAPDH. N = 9 (a) or 7 (b) independent experiments. c HIF1α Immunoprecipitation (IP) or control IP (Ctrl) and HIF1α Western blot in lysates of rat SCs cultured as above (a) and incubated with MG-132 for 8 h. Same lysate separated into two equal fractions (for HIF1α IP and Ctrl IP). GAPDH input=3% of IP lysate. Three independent experiments, representative images are shown. d IP HIF1α or Ctrl IP and TRAF7 Western blot in lysates of rat SCs cultured as above (a) and incubated for 8 h with MG-132. Three independent experiments, representative images are shown. Same lysate separated into two equal fractions, (for HIF1α IP and Ctrl IP). TRAF7 and GAPDH inputs=3% of IP lysate. e Denaturing HIF1α and Ctrl IP and ubiquitin (P4D1) Western blot in lysates of SCs transduced with T7sh or Csh lentivirus cultured as above (a) and incubated for 8 h with MG-132, and quantification of ubiquitinated HIF1α. N = 3 independent experiments. Each lysate separated into two equal fractions (for HIF1α IP and Ctrl IP). GAPDH input=3% of IP lysate. f TRAF7 Western blot on lysates of rat SCs transfected with a TRAF7- or Flag-expressing construct (Ctrl), and quantification normalized to GAPDH. N = 3 independent experiments. g–i HIF1α (g), c-Jun (h) or phospho-c-Jun (p-c-Jun, i) Western blots on lysates of rat SCs cultured as above (b) and incubated with lentiviruses carrying an HDAC8 shRNA (H8sh) or Csh, and subsequently transfected with a TRAF7- or Flag-expressing construct (Ctrl), and quantification normalized to GAPDH. N = 3 independent experiments. Paired (a, b, e, g, h, i/Ctrl) or unpaired (f, i/Csh-TRAF7) two-tailed (black asterisks) or one-tailed (gray asterisks, n.s.) Student’s t-tests, p value: *<0.05, **<0.01, n.s. = non-significant, values = mean, error bars = s.e.m. Source data are provided as a Source data file.
