Expression data from mouse wild type, PRDM14-overexpressing and Dnmts KO embryonic stem cells (ESCs).. Mus musculus

Germline cells reprogram extensive epigenetic modifications to ensure the cellular totipotency of the next generation and prevent accumulation of epimutations. Primordial germ cells (PGCs)1, the common source of both oocytes and sperm, erase genome-wide DNA methylation and histone H3 lysine 9 dimethylation (H3K9me2), a process called genome-wide epigenetic reprogramming2,3. However, little is known about the molecular mechanism of DNA demethylation by developing PGCs. Here we show that overexpression of PRDM14, a critical regulator for specification and early differentiation of PGCs, promotes global DNA demethylation in embryonic stem cells (ESCs). PRDM14 directly represses transcription of de novo DNA methyltransferase, Dnmt3b, but its repression is not sufficient for global DNA demethylation. Comparison of global gene expression profiles between PRDM14-overexpressing ESCs and Dnmts triple mutant ESCs clearly demonstrates that overexpression of PRDM14 activates about half of the genes silenced by DNA methylation in ESCs. Furthermore, PRDM14 directly interacts with TET1, which converts 5-methylcytosine to 5-hydroxymethylcytosine, and DNA demethylation by overexpression of PRDM14 is strongly disturbed by pharmacological inhibitors of the base excision repair (BER) pathway. We propose that formation of a PRDM14/TET1 complex triggers the activation of BER-dependent active demethylation across the whole genome of developing PGCs. Overall design: To determine the effects of DNA demethylation by overexpression of PRDM14 on the global gene expression profile, we exploited microarray analysis using total mRNAs derived from J1 (control), PRDM14-overexpressing, PRDM14-overexpressing + DNMT3B1, Dnmt3b KO, Dnmt1 KO and Dnmts TKO mouse ES cells, in total 10 samples.