Research Article

Structure of human TFIID and mechanism of TBP loading onto promoter DNA

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Science  21 Dec 2018:
Vol. 362, Issue 6421, eaau8872
DOI: 10.1126/science.aau8872

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Mechanism of promoter recognition

To start transcription, RNA polymerase II is recruited by the general transcription factor IID (TFIID) to the DNA promoter. Patel et al. used a combination of experimental approaches to elucidate the full molecular architecture of human TFIID and its complete conformational landscape during promoter recognition. They suggest exactly how TFIID is loaded onto the promoter, which involves defined steps—including promoter recognition and transcription initiation—and leads to regulated gene expression.

Science, this issue p. eaau8872

Structured Abstract

INTRODUCTION

In eukaryotes, transcription initiation starts with the assembly of the transcription preinitiation complex (PIC) onto promoter DNA. The PIC comprises the general transcription factors and RNA polymerase II (Pol II). The general transcription factor IID (TFIID) is responsible for initially recognizing the core promoter. Human TFIID is a trilobed (lobes A, B, and C) complex composed of TATA box binding–protein (TBP) and 13 evolutionarily conserved TBP-associated factors (TAF1 to TAF13), with six TAFs present in two copies. Together, TBP and the TAF subunits of TFIID directly interact with promoter DNA with the assistance of TFIIA, forming a platform for the assembly of the rest of the PIC.

RATIONALE

A key challenge in understanding the molecular basis behind TFIID’s recognition of promoter DNA is the lack of a complete structural depiction of the complex. We used cryo–electron microscopy (cryo-EM) to describe the various biochemical and/or conformational states of the complex, thus providing information on both the structure and dynamics of TFIID and its interaction with promoter DNA.

RESULTS

We report the cryo-EM structure of TFIID with a resolution of 4.3 Å for lobe C, 4.5 Å for lobe B, and 9.8 Å for lobe A. Together with chemical cross-linking mass spectrometry and structure prediction, we generated a complete structural model of the evolutionarily conserved core of TFIID. TFIID is built on a dimeric scaffold of TAFs, containing at its center a TAF6 dimer in lobe C that connects to lobes A and B. Lobes A and B are both organized around TAF4, -5, -6, -9, -10, and -12 but include additional subunits that result in distinct function (see the figure). Lobe A, which contains TAF11 and TAF13 interacting with TBP, keeps TBP inhibited unless TFIID is promoter bound, at which point it loads TBP onto DNA. Lobe B contains TAF8, which extends to hold lobes B and C together rigidly. Lobe B positions TAF4 in place to stabilize upstream DNA binding and recruits TFIIA. Lobe C, in addition to TAF6 and TAF8, contains TAF1, -2, and -7, which bind the downstream core promoter sequences.

Using computational sorting of cryo-EM images, we characterized the conformational landscape of apo-TFIID and TFIID in the presence of TFIIA and promoter DNA. Two major states for apo-TFIID (termed the canonical and extended states) were observed, and three additional states (termed the scanning, rearranged, and engaged states) were observed in the presence of TFIIA and core promoter DNA. Lobe A, which migrates 150 Å from its position near lobe C in the canonical state to near lobe B in the extended state, carries TBP in a repressed state that is only released in the context of promoter binding. Identification of distinct TFIID states allowed us to generate a mechanistic model for TFIID promoter binding (see the figure). We propose that TFIID first binds the downstream core promoter elements through TAF1 and TAF2. This binding and the flexible attachment of lobe A help position the upstream DNA in proximity to TBP. TBP then scans for a TATA box or its sequence variants. Engagement of upstream core promoter sequences by TBP is facilitated by TFIIA interacting with TAF4 and TAF12 within lobe B. When TBP finally binds the promoter, it releases from lobe A, opening the binding site for TFIIB, which can then recruit Pol II.

The structure of TFIID also allowed us to deduce the position of various regulatory domains of TFIID involved in contacts with transcriptional activators and active chromatin marks that are responsible for recruiting and modulating TFIID function.

CONCLUSION

Our studies lead to a mechanistic model of how TFIID prevents TBP from nonspecifically engaging with DNA outside of gene promoters, thus preventing aberrant PIC assembly and erroneous transcription initiation. Our model also suggests how TFIID loads TBP onto TATA-less promoters and how activators and chromatin marks may direct TFIID recruitment and PIC assembly.

Structure of human TFIID.

The structure of apo-TFIID is shown in the canonical state and that of promoter-bound TFIID is depicted in the engaged state. Lobes A and B in TFIID share a similar architecture that contains histone-fold domains organized in a manner that resembles a histone octamer. The TAF6 subunit of TFIID dimerizes the core set of TAFs. The TAF8 subunit rigidly tethers together lobes B and C. Five states of TFIID were observed in the process of promoter binding, leading to a mechanistic model of TBP loading onto the promoter DNA.

Abstract

The general transcription factor IID (TFIID) is a critical component of the eukaryotic transcription preinitiation complex (PIC) and is responsible for recognizing the core promoter DNA and initiating PIC assembly. We used cryo–electron microscopy, chemical cross-linking mass spectrometry, and biochemical reconstitution to determine the complete molecular architecture of TFIID and define the conformational landscape of TFIID in the process of TATA box–binding protein (TBP) loading onto promoter DNA. Our structural analysis revealed five structural states of TFIID in the presence of TFIIA and promoter DNA, showing that the initial binding of TFIID to the downstream promoter positions the upstream DNA and facilitates scanning of TBP for a TATA box and the subsequent engagement of the promoter. Our findings provide a mechanistic model for the specific loading of TBP by TFIID onto the promoter.

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