The complete structure of the human TFIIH core complex.

Basil J Greber,Daniel B Toso,Jie Fang,Eva Nogales

doi:10.7554/elife.44771

Abstract

Transcription factor IIH (TFIIH) is a heterodecameric protein complex critical for transcription initiation by RNA polymerase II and nucleotide excision DNA repair. The TFIIH core complex is sufficient for its repair functions and harbors the XPB and XPD DNA-dependent ATPase/helicase subunits, which are affected by human disease mutations. Transcription initiation additionally requires the CdK activating kinase subcomplex. Previous structural work has provided only partial insight into the architecture of TFIIH and its interactions within transcription pre-initiation complexes. Here, we present the complete structure of the human TFIIH core complex, determined by phase-plate cryo-electron microscopy at 3.7 Å resolution. The structure uncovers the molecular basis of TFIIH assembly, revealing how the recruitment of XPB by p52 depends on a pseudo-symmetric dimer of homologous domains in these two proteins. The structure also suggests a function for p62 in the regulation of XPD, and allows the mapping of previously unresolved human disease mutations.

Highlights

Transcription factor IIH (TFIIH) is a 10-subunit protein complex with a total molecular weight of 0.5 MDa that serves a dual role as a general transcription factor for transcription initiation by eukaryotic RNA polymerase II (Pol II), and as a DNA helicase complex in nucleotide excision DNA repair (NER) (Compe and Egly, 2016; Sainsbury et al, 2015)
To determine the complete structure of the human TFIIH core complex, we collected several large cryo-electron microscopy (cryo-EM) datasets (Supplementary file 1) of TFIIH immuno-purified from HeLa cells using an electron microscope equipped with a Volta phase plate (VPP) (Danev and Baumeister, 2017) and a direct electron detector camera mounted behind an energy filter
This VPP-based cryo-EM map was substantially improved compared to our previous maps obtained without phase plate, both in resolution and interpretability (Figure 1—figure supplement 2D–G), and enabled building, refinement, and full validation of an atomic model of the TFIIH core complex and the MAT1 subunit of the CAK subcomplex (Figure 1A–C, Figure 1—figure supplement 2B,C, Supplementary file 2, 3), while the remainder of the CAK subcomplex is invisible in our map because it is flexibly tethered to the TFIIH core complex

Summary

Introduction

Transcription factor IIH (TFIIH) is a 10-subunit protein complex with a total molecular weight of 0.5 MDa that serves a dual role as a general transcription factor for transcription initiation by eukaryotic RNA polymerase II (Pol II), and as a DNA helicase complex in nucleotide excision DNA repair (NER) (Compe and Egly, 2016; Sainsbury et al, 2015). The TFIIH core complex is composed of the seven subunits XPB, XPD, p62, p52, p44, p34, and p8, and is the form of TFIIH active in DNA repair (Svejstrup et al, 1995), where TFIIH serves as a DNA damage verification factor (Li et al, 2015; Mathieu et al, 2013) and is responsible for opening a repair bubble around damaged nucleotides This activity depends on both the SF2-family DNA-dependent ATPase XPB, and the DNA helicase activity of XPD (Coin et al, 2007; Evans et al, 1997; Kuper et al, 2014).

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Results

Conclusion