mNET-seq on COMPASS-degron cells

We measured the the dwell time of existed paused RNAPII at the promoter by mammalian native elongating transcript sequencing (mNET-seq). We found a median RNAPII half-life of 5.3 min at protein-coding genes in DMSO pre-treated control mESCs, whereas the RNAPII half-life at these genes significantly slowdown to 8.96 min when pre-treated with Auxin in the DPY30-mAID degron cell. The data suggested that loss H3K4me3 directly slows down the release of paused RNAPII throughout genes. To directly investigate the temporal stability and half-life of promoter RNAPII, we used Triptolide, a potent inhibitor of transcription initiation, coupled with mammalian native elongating transcript sequencing (mNET-seq) to measure the dwell time of existed paused RNAPII at the promoter. Unlike previous measurements, the single-nucleotide resolution of the mNET-seq data allowed us to specifically direct measure the RNAPII engaged RNA signal at protomer-proximal region.