Role of Tet proteins in enhancer activity and telomere elongation

Abstract

DNA methylation at the C-5 position of cytosine (5mC) is one of the best-studied epigenetic modifications and plays important roles in diverse biological processes. Iterative oxidation of 5mC by the ten-eleven translocation (Tet) family of proteins generates 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). 5fC and 5caC are selectively recognized and excised by thymine DNA glycosylase (TDG), leading to DNA demethylation. Functional characterization of Tet proteins has been complicated by the redundancy between the three family members. Using CRISPR/Cas9 technology, we generated mouse embryonic stem cells (ESCs) deficient for all three Tet proteins (Tet triple knockout [TKO]). Whole-genome bisulfite sequencing (WGBS) analysis revealed that Tet-mediated DNA demethylation mainly occurs at distally located enhancers and fine-tunes the transcription of genes associated with these regions. Functional characterization of Tet TKO ESCs revealed a role for Tet proteins in regulating the twocell embryo (2C)-like state under ESC culture conditions. In addition, Tet TKO ESCs exhibited increased telomeresister chromatid exchange and elongated telomeres. Collectively, our study reveals a role for Tet proteins in not only DNA demethylation at enhancers but also regulating the 2C-like state and telomere homeostasis.

Tet1 and Tet2 Protect DNA Methylation Canyons against Hypermethylation

“…Our unbiased analysis of genome-wide methylation patterns showed widespread hypermethylation in DKO MEFs, which included promoter and enhancer regions. These findings are consistent with recent studies of Tet-deficient mouse ESCs (11,12) and provide a mechanistic explanation for the epigenetic regulatory defects of Tet-deficient cells.…”

“…S2 in the supplemental material). Significant (P Ͻ 0.01; Wilcoxon rank sum test) hypermethylation was detected for both promoter and enhancer regions, consistent with recent findings in Tet-deficient mouse ESCs (11,12). As Tet-dependent methylation changes at promoters appeared more prominent (Fig.…”

“…DKO mice are characterized by global DNA hypermethylation and developmental plasticity, suggesting an important role of Tet1/Tet2 in the epigenetic regulation of developmental genes (8). In agreement with this notion, recent studies have shown a role of Tet-mediated demethylation in the modulation of enhancer activity (10)(11)(12).…”

“…P values to indicate significant differential expression for the transcriptome sequencing (RNA-seq) analysis were obtained from DESeq after FDR correction (27). Statistical testing for the overlap between differentially expressed genes and genes associated with hypermethylated canyons was done by a hypergeometric test using R, as described previously (11). Biological pathway analyses were performed using Ingenuity Pathway Analysis software, and bar charts showing the Ϫlog (P values) were directly used as figure panels.…”

Epigenetic Memory Underlies Cell-Autonomous Heterogeneous Behavior of Hematopoietic Stem Cells

“…To evaluate the sensitivity of our CNV calling method for bisulfite whole genome sequencing data, we analyzed published bisulfite whole genome sequencing data for two different mouse strains: 129P2 (GSE56986) (Lu et al, 2014) and BALBcJ (GSE60485) (Wu et al, 2015). The CNVs for the three mouse strains were annotated by Sanger mouse genome project.…”