Stress-Induced Transcriptional Memory Accelerates Promoter-Proximal Pause-Release and Decelerates Termination over Mitotic Divisions

Anniina Vihervaara; Dig Bijay Mahat; Samu V. Himanen; Malin A.H. Blom; John T. Lis; Lea Sistonen

To address whether heat-induced reprogramming of RNA synthesis is followed by restoration or 106! readjustment of transcription, we measured nascent RNA synthesis in MEFs upon a 1-hour heat 107! shock, and after a 4-hour or a 48-hour recovery (Figure 1 ). DNA staining and Fluorescence 108! Associated Cell Sorting (FACS) verified that MEFs proliferated over the acute heat shock and 109! also following recovery, and they did not undergo cell cycle arrest or apoptosis ( Figure S1A ). For 110! accurate normalization of PRO-seq data, we used whole-genome spike-in from Drosophila S2 111! cells ( Figure S1B , see Materials and Methods for details), and pooled biological replicates for 112! data analyses after validating their high correlation ( Figure S1C ). As previously reported (Mahat 113! et al., 2016; , a single heat shock caused a robust induction of hundreds 114! and repression of thousands of genes, simultaneously accumulating Pol II at divergently 115! transcribed distal Transcription Regulatory Elements (dTREs), hereon called enhancers (Figure 116! 1A). During a 4-hour recovery, the genome-wide profile of gene and enhancer transcription was 117! precisely restored to the level observed prior to the heat shock ( Figure 1A -B), which demarcates 118! high plasticity and accurate recovery of the transcription program. Despite the full recovery of 119! gene body transcription, certain promoter-proximal regions gained new pause sites, as 120! exemplified by Heat Shock Protein H1 (Hsph1 alias Hsp110) gene ( Figure 1B ). Moreover, a 121! subset of heat-responsive genes either gained ( Figure S1D ) or retained ( Figure S1E ) high Pol II 122! density at a single pause site during the recovery. Consequently, the genome-wide average of Pol 123! II at the pause region remained elevated, even when measured in the daughter cells, 48 hours 124! after the heat exposure ( Figure 1C ). 125!