Individual retrotransposon integrants are differentially controlled by KZFP/KAP1-dependent histone methylation, DNA methylation and TET-mediated hydroxymethylation in naïve embryonic stem cells. Mus musculus

The KZFP/KAP1 (KRAB zinc finger proteins/KRAB-associated protein 1) system plays a central role in the silencing of transposable elements (TEs) and the maintenance of parent-of-origin DNA methylation at imprinting control regions (ICRs) during the wave of genome-wide reprogramming that precedes implantation. In naïve murine embryonic stem cells (mESCs), the genome is maintained highly hypomethylated by a combination of active demethylation (operated by TET proteins) and lack of de novo methylation; in these cells, KAP1 is tethered by sequence-specific KZFPs to ICRs and TEs where it recruits histone and DNA methyltransferases to impose heterochromatin formation and DNA methylation. Here, upon removing either KAP1 or the cognate KZFP, we observed rapid TET-dependent accumulation of 5hmC at both ICRs and TEs. In absence of the KZFP/KAP1 complex, ICRs lost heterochromatic histone marks and underwent both active and passive DNA demethylation. Using RNA-seq, we further compared the expression profiles of TEs upon Kap1 removal in wild type, Dnmt and Tet triple knockout mESCs. We found that KAP1 represents the main effector of TEs repression in all three settings, yet we could additionally identify specific groups of TEs also controlled by DNA methylation. Furthermore, activation upon Kap1 removal in the absence of TET proteins could be either blunted or increased depending on TE subsets, indicating a complex interplay between heterochomatin and active demethylation in regulating the expression of the endovirome. Overall design: Profile of H3K4me3 and H3K27me3 in naive murine emrbyonic stem cells Profile of 5-hydroxymethylcytosine enrichment in mure embryonic stem cells upon Kap1 knockdown Analysis of transcriptional profiles of wild type, Dnmt triple knockout and Tet triple knockout murine embryonic stem cells upon Kap1 knockdown