Fig. 4

Epigenetic control of Th and Nurr1 gene transcription by JMJD3. (A) Real-time PCR analysis of mRNA levels of Jmjd3, Th and Nurr1 from striatum (STRM) and midbrain (MB) in Jmjd3 conditional knockout mice (cKO) and their control littermates (WT). N = 3 mice. Data represent mean ± SEM. (B) Representative immunofluorescence images and quantifications of NURR1 expression (green) in tyrosine hydroxylase (TH; red) + cells from primary culture of mouse midbrain dopamine (mDA) neurons with or without treatment of GSK-J4. Insets show cell nuclei. Scale bar represents 20 μm. (C) Real-time PCR analysis of Th and Nurr1 mRNA levels in primary cultured mDA neurons after GSK-J4 treatment. (D) Representative immunofluorescence images of histone 3 lysine 27 tri-methylation (H3K27me3) in TH + mDA neurons from cKO, GSK-J4 mice, Svct2 KO mouse embryos and primary cultured mDA neurons treated with GSK-J4. Scale bar represents 20 μm. (E, F) Chromatin immunoprecipitation (ChIP)-quantitative PCR analysis of H3K27me3, JMJD3 and NURR1enrichments on 1 kilo bp mouse Th (E) and Nurr1 (F) promoters from DA neuronal cell line MES23.5 with or without GSK-J4 treatment. Blue boxes represent consensus NURR1 binding sites. Nd = not detected. N = 3 independent cell cultures. Data represent mean ± SEM. *P < 0.05; Student’s t-test and one-way ANOVA with Tukey’s post-hoc test