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Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB, and XPB.

ABSTRACT: General transcription factors (GTFs) are required for RNA polymerase II (Pol II) to initiate transcription at promoters. In this study, we determined the effects of acute depletion of TBP, TAF1, TAF4, TFIIB, and XPB in HAP1 cells. We performed precision nuclear run-on sequencing (PRO-Seq) and quantified nascent transcripts arising from more than 70,000 promoters. The average dependencies for each factor across all promoters varied widely even though levels of depletions were similar. Many of the effects could be attributed to the presence or absence of core promoter elements. Depletion of TBP had a large effect on only a small fraction of Pol II and Pol III promoters. TFIIB depletion also led to readthrough transcription downstream of the 3′ ends of genes. We conclude that promoter activity is influenced by recruitment of TFIID, sequence-specific transcription factors, and interaction of the preinitiation complex (PIC) with the +1 nucleosome.

ORGANISM(S): Homo sapiens

PROVIDER: GSE194153 | GEO | 2022/07/17

REPOSITORIES: GEO

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Project description:RNA polymerase II (Pol II) transcription initiation starts with the assembly of the preinitiation complex (PIC) on core promoters. The PIC is composed of six general transcription factors (GTFs). The recognition of the core promoter sequences by the TFIID GTF complex is the first step of the PIC assembly. In metazoans, holo-TFIID is composed of the TATA binding protein (TBP) and of 13 TBP associated factors (TAFs). Genetic depletion of different murine TAFs have shown that TAFs such as TAF7 or TAF10, can be either required, or dispensable for Pol II transcription, depending on different cellular contexts. In this report, we depleted TAF7 and/or TAF10 in the same cellular system; either in mesodermal progenitors during mouse development or in mESCs. In these two models, TAF7 depletion leads to a milder phenotype compared to TAF10 depletion. As TAF10 is also a subunit of the transcriptional co-activator Spt-Ada-Gcn5 acetyl transferase (SAGA), we first showed that the difference in phenotype between the Taf7 and Taf10 mutant is not due to the SAGA effect, at least for mESCs. Immunoprecipitations coupled with mass spectrometry analyses from mESCs lysates assembly of holo-TFIID complex is rapidly affected after induction of the depletion. In line with the model of holo-TFIID sequential assembly, TAF10 depletion leads to an early defect with formation of the core-TFIID, while TAF7 depletion results in the formation of a TAF7-less TFIID. Thus, the difference in phenotype severity correlates with the degree of TFIID disassembly. Surprisingly, no major global changes in Pol II transcription could be observed after either TAF7 or TAF10 depletion. Our data suggest that the inducible loss of fully assembled canonical TFIID does not correlate with the lack of global Pol II transcription activity changes suggesting that partially assembled TFIID complexes can participate in Pol II transcription initiation, with only limited effect on Pol II nascent transcription.

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Differential dependencies of human RNA polymerase II promoters on TBP, TAF1, TFIIB, and XPB.

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