Abstract
A repertoire of transcription initiation factors engage the core promoter of mRNA genes to recruit RNA polymerase (Pol) II to initiate transcription, yet their precise spatial organization remains unclear. Using ChIP-exo, here we detail the interactions and genomic organization of initiation factors TBP, TFIIB, and Pol II at mRNA genes and within CpG islands. We find that when Pol II moves into a transcriptionally paused state, TBP/TFIIB remain at the promoter. We show that TBP and TFIIB bound to the core promoter at two separate, resolvable locations that coincided with sites of divergent transcription initiation. We also examine the precise binding of TBP at Pol III transcribed tRNA genes. We find that TBP crosslinked to tRNA genes in a similar manner as at Pol II transcribed genes. This comprehensive and high resolution genome-wide detection of the initiation machinery produces a consolidated view of transcription initiation events humans at Pol II coding and Pol III transcribed tRNA genes.
Highlights
The classic paradigm for assembling the minimal core transcription machinery at mRNA promoters starts with the recruitment of the TATA binding protein (TBP)
We focused on TBP and TFIIB to assess Pre-initiation complex occupancy (PIC) formation because in yeast these proteins were the most detail-rich, whereas other initiation factors displayed essentially similar ChIP-exo patterns [15]
To assess post-initiation transcription complexes and the extent to which genes display promoter-proximal pausing, we ChIP’d the largest Pol II subunit (POLR2A). 8,364 TFIIB ChIP-exo peak-pairs (Table A in S1 File) were found within 500 bp of an mRNA TSS, which corresponds to ~50% of all annotated protein-coding K562-expressed genes (Fig 1A)
Summary
The classic paradigm for assembling the minimal core transcription machinery at mRNA promoters starts with the recruitment of the TATA binding protein (TBP). Together with TFIIF, TFIIB engages Pol II in its active site to help set the start site of transcription (TSS) [1, 2]. Further complicating the classic paradigm of transcription initiation of mRNA genes is the coupling of antisense transcription upstream of the core promoter [8]. These divergent TSSs are spaced roughly 250 bp apart with some variance, and driven by separate initiation complexes [9]. The precise genomic organization of human transcription complexes within this context remains
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