Fig. 3

Kdm3a and Kdm3b mediate vitamin C-induced loss of H3K9me2. a Kinetics and reversibility of H3K9me2 loss following vitamin C treatment. H3K9me2 levels were measured by western blot at 6, 24, and 72 h following vitamin C treatment. Vitamin C was then removed, and H3K9me2 levels were measured 72 h later (144 h). b Western blot for H3K9me2 in ES cells ± vitamin C with siRNA knockdown of Kdm3 family enzymes or a non-targeting control. c Western blot for H3K9me2 in ES cells ± vitamin C with double knockdown of Kdm3a/3b or non-targeting control. d ChIP-qPCR for H3K9me2 at gene promoters in ES cells ± vitamin C with double knockdown of Kdm3a/3b or non-targeting control. Data are presented as fold change relative to the untreated control. Data are mean ± SD. Asterisks represent P < 0.05 by t test. e In vitro activity of recombinant KDM3A toward demethylation of a synthetic H3K9me1 peptide, in the presence of vitamin C, DTT or glutathione (see “Methods” section for details). Data are mean ± SD. Asterisks are P < 0.05 by t test for vitamin C compared to the oxidized form of glutathione