TET2 Loss Triggers ssDNA Gap Accumulation and Heightened PARP Inhibitor Sensitivity

Here we sought to understand the molecular effects of TET2 knock out (KO) in U2OS cells. TET2 catalyzes the conversion of 5-methyl-cytosine to 5-hydroxy-methyl-cytosine (5hmC). We thus mapped 5hmC genome-wide using bACE-seq (bisulfite APOBEC-coupled epigenetic sequencing) in TET2 wild-type (WT) and KO. TET2 KO U2OS cells were engineered to remove ~42kb of TET2. TET2 KO was confirmed by PCR, western blot, and sequencing. bACE-seq analysis was then performed on genomic DNA isolated from three independent plates of WT and KO U2OS cells.

Our subsequent analysis identified 4182 differentially hypo-hydroxy-methylated regions (hypo-hDMRs) and 651 hyper-hydroxy-methylated DMRs, confirming an overall reduction in the genomic levels of cytosine hydroxy-methylation upon TET2 loss. Hypo-hDMRs were spread across the chromosomes, and we did not observe specific chromosomal regions having preferential hDMR concentration. Moreover, we did not observe localization of hDMRs to any specific genomic compartment such as promoters, enhancers, or UTRs. Using previously published replication initiation and termination zones defined by OK-seq we observed that in WT cells initiation zones showed a slight reduction in 5hmC levels relative to randomized genomic control regions and termination zones. In TET2 KO cells, the difference at initiation zones was dampened as likely all genomic regions have reduced 5hmC. Here we provide bACE-seq hmC mapping data for TET2 WT and KO U2OS cells for three replicates of each condition.

• Type: Methylation Sequencing
• Archive: The database of Genotypes and Phenotypes (dbGaP)