The Histone H3 Lysine 27-Specific Demethylase Jmjd3 Is Required for Neural Commitment (original) (raw)
Figure 4
Jmjd3 regulates key effectors of neurogenesis.
(a) Loss of Jmjd3 impairs upregulation of the neuronal markers Pax6, Nestin and Sox1 during differentiation. Levels of Pax6, Nestin and Sox1 were quantified by real-time RT-PCR. Bars represent the means±s.d. of triplicates normalized to TBP. W.t.: wild-type ES cells; Jmjd3-kd: Jmjd3 knock-down cells. (b) Scheme of the genomic regions analyzed with chromatin immunoprecipitation. The Pax6 (upper panel), Nestin (middle panel) and Sox1 (bottom panel) loci are drawn to scale, showing (partial) exon-intron structure, the transcription start site(s), the region covered by the so-called bivalent domains (see text) and the amplicons analyzed by qPCR after immunoprecipitation. (c) Chromatin immunoprecipitation showing levels of Jmjd3 occupancy of the genomic regions outlined in 3b in normal ES cells during differentiation. Levels of enrichment are shown as percentage of input chromatin. Error bars represent standard deviations of qPCR triplicates. undiff.: undifferentiated ES cells; day4 and day8 represent two time points of the differentiation protocol. (d) Chromatin immunoprecipitation showing levels of H3-K27me3 on the genomic regions outlined in 3b in normal ES cells during differentiation. Levels of enrichment are shown as percentage of input chromatin. Bars are the average +/−S.E.M. of triplicate independent samples. Undiff.: undifferentiated ES cells; day8: time point after the beginning of the differentiation protocol; NSC: neural stem cells (day 26). (e) Chromatin immunoprecipitation showing levels of H3-K27me3 on the Nestin promoter (outlined in 3b) in wild type (w.t.) and Jmjd3-kd (Jmjd3-kd1 and 2) cells in the undifferentiated state (undiff.) and at day 7 of monolayer differentiation. Levels of enrichment are shown as percentage of input chromatin. Bars are the average +/−S.E.M. of triplicate independent samples.