LINE1 and PRC2 control nucleolar organization and repression of the 8C-state in human ESCs [ChIRP-seq,RSeT+DT H9]

In this study, we profiled for LINE1 binding loci in RSeT+DT naïve hES cells with the Chromatin Isolation by RNA Purification (ChIRP)-seq strategy. We followed a previously published ChIRP protocol described (Percharde et al. 2018; Lu et al. 2020). Overall design: RSeT H9 hES cells were witched to RSeT media added with 10 nM DZNep and 5nM TSA (RSeT+DT) for 48 hours, then collected for preparing ChIRP sequencing sample. x107 cells per ChIRP pulldown were subjected to DNAseI digestion after crosslinking the cells. After brief sonication and spinning, chromatin supernatants were collected for hybridization with probes. After 3 hour of hybridization with LINE1 probes or the control MALAT1 probes, streptavidin M280 beads were added and incubated for another 3 hours. After hybridization, the beads were washed 5 times and then treated with RNase H to elute ChIRP pulled down DNAs, which were reverse crosslinked and purified, and then used for library preparation. More protocol details are described in the methods of this study.