Human RNA polymerase III transcriptomes and relationships to Pol II promoter chromatin and enhancer-binding factors

Abstract

RNA polymerase (Pol) III transcribes many noncoding RNAs (e.g. tRNAs) important for translational capacity and other functions. Here, we localized Pol III, alternative TFIIIB complexes (BRF1/2) and TFIIIC in HeLa cells, determining the Pol III transcriptome, defining gene classes, and revealing ‘TFIIIC-only’ sites. Pol III localization in other transformed and primary cell lines revealed novel and cell-type specific Pol III loci, and one miRNA. Surprisingly, only a fraction of the in silico-predicted Pol III loci are occupied. Many occupied Pol III genes reside within an annotated Pol II promoter. Outside of Pol II promoters, occupied Pol III genes overlap with enhancer-like chromatin and enhancer-binding proteins such as ETS1 and STAT1. Remarkably, Pol III occupancy scales with the levels of nearby Pol II, active chromatin and CpG content. Taken together, active chromatin appears to gate Pol III accessibility to the genome.

RNA Polymerase III Accurately Initiates Transcription from RNA Polymerase II Promoters in Vitro

“…Genome-wide analysis of Pol III promoter occupancy in HeLa cells suggested promoter accessibility to be a major regulator of Pol III transcription (53). This was also observed in vitro.…”

High-resolution mapping of transcriptional dynamics across tissue development reveals a stable mRNA–tRNA interface

“…For each experiment at each tissue and developmental stage, we performed two biological replicates that were highly correlated Methods). This approach allowed us to quantify expression levels for protein-coding genes, as well as Pol III occupancy at every tRNA locus, which quantitatively captures the utilization of each tRNA gene (Barski et al 2010;Moqtaderi et al 2010;Oler et al 2010;Kutter et al 2011;Canella et al 2012;Carriere et al 2012;Renaud et al 2014).…”

“…Previous studies reported that active chromatin coincides with active transcription of Pol III genes (Barski et al 2010;Oler et al 2010). We therefore used previously published data (Shen et al 2012) to compare the occurrence of three histone marks associated with transcriptionally active chromatin modifications, histone H3 lysine 27 acetylation (H3K27ac), H3 lysine 4 tri-(H3K4me3) and monomethylation (H3K4me1), Pol II, and the insulator-binding protein CCCTC-binding factor (CTCF) at genomic localizations 0.1, 0.5, and 1.0 kb near differentially expressed tRNA genes (Shen et al 2012;Methods).…”

“…We did this since prior studies have shown that sequence variation in internal regulatory sequences of tRNA genes have no clear relationship with their expression levels (Oler et al 2010;Canella et al 2012). We searched for regulatory sequence elements acting in cis that could direct the Pol III recruitment to tRNA genes as described previously (Giuliodori et al 2003;Bloom-Ackermann et al 2014).…”

“…gene usage and transcript abundance (Methods) (Barski et al 2010;Moqtaderi et al 2010;Oler et al 2010;Kutter et al 2011;Canella et al 2012;Carriere et al 2012;Renaud et al 2014). From E15.5 to P29 in liver and brain, 311 of the 433 tRNA genes predicted by tRNAscan-SE (Schattner et al 2005) were utilized (Supplemental Fig.…”