Jmjd2a and Jmjd2c regulate H3K9me3 and H3K36me3 at H3K4me3 positive transcription start sites being essential for ESC self-renewal and embryogenesis (ChIP-Seq). Mus musculus

We have characterized the role of the Jmjd2/Kdm4 proteins in embryonic stem cell (ESC) biology, histone methylation and gene regulation. The Jmjd2 proteins are H3K9/H3K36 histone demethylases and three Jmjd2 family members are expressed in ESCs: Jmjd2a/Kdm4a, Jmjd2b/Kdm4b and Jmjd2c/Kdm4c/Gasc1. We find that specifically Jmjd2a and Jmjd2c exert redundant functions, which are essential for ESC self-renewal and early embryonic development. ChIP-seq studies show that Jmjd2a and Jmjd2c both localize to H3K4me3 marked regions, where they have general and widespread roles preventing the accumulation of especially H3K9me3, but also H3K36me3. Jmjd2 catalytic activity is required for ESC maintenance, and increased H3K9me3 levels in knockout ESCs compromise the expression of several Jmjd2a/c targets, including genes that are important for ESC self-renewal. Thus, continual removal of H3K9 promoter methylation by Jmjd2 demethylases represents a novel mechanism ensuring transcriptional competence and stability of the pluripotent cell identity. Overall design: For the identification of Jmjd2a binding patterns, ChIP experiments were performed using conditional Jmjd2a knockout ESCs (Jmjd2a(f/f);Rosa26::CreERT2). The OHT-treated sample (devoid of Jmjd2a) serves as a negative control. H3K9me3, H3K36me3, H3K4me3 and H3 distributions were mapped for conditional Jmjd2a/c double (Jmjd2a(f/f);Jmjd2c(f/f);Rosa26::CreERT2/CreERT2) and Jmjd2a/b/c triple (Jmjd2a(f/f);Jmjd2b(f/f);Jmjd2c(f/f);Rosa26::CreERT2/CreERT2) knockout ESCs grown in the absence or presence of OHT to induce loss of Jmjd2 expression. ESCs were cultured in 2i serum-free medium.