TFIID TAF6-TAF9 Complex Formation Involves the HEAT Repeat-containing C-terminal Domain of TAF6 and Is Modulated by TAF5 Protein

Elisabeth Scheer,Frédéric Delbac,Laszlo Tora,Dino Moras,Christophe Romier

doi:10.1074/jbc.m112.379206

Abstract

The general transcription factor TFIID recognizes specifically the core promoter of genes transcribed by eukaryotic RNA polymerase II, nucleating the assembly of the preinitiation complex at the transcription start site. However, the understanding in molecular terms of TFIID assembly and function remains poorly understood. Histone fold motifs have been shown to be extremely important for the heterodimerization of many TFIID subunits. However, these subunits display several evolutionary conserved noncanonical features when compared with histones, including additional regions whose role is unknown. Here we show that the conserved additional C-terminal region of TFIID subunit TAF6 can be divided into two domains: a small middle domain (TAF6M) and a large C-terminal domain (TAF6C). Our crystal structure of the TAF6C domain from Antonospora locustae at 1.9 Å resolution reveals the presence of five conserved HEAT repeats. Based on these data, we designed several mutants that were introduced into full-length human TAF6. Surprisingly, the mutants affect the interaction between TAF6 and TAF9, suggesting that the formation of the complex between these two TFIID subunits do not only depend on their histone fold motifs. In addition, the same mutants affect even more strongly the interaction between TAF6 and TAF9 in the context of a TAF5-TAF6-TAF9 complex. Expression of these mutants in HeLa cells reveals that most of them are unstable, suggesting their poor incorporation within endogenous TFIID. Taken together, our results suggest that the conserved additional domains in histone fold-containing subunits of TFIID and of co-activator SAGA are important for the assembly of these complexes.

Highlights

TFIID/SAGA histone fold-containing subunits contain conserved additional domains of unknown function
Our results suggest that the conserved additional domains in histone fold-containing subunits of TFIID and of co-activator SAGA are important for the assembly of these complexes
Numerous biochemical and structural studies have shown that the histone fold motif plays a major role in TFIID and SAGA assembly, enabling the formation of five distinct histone-like pairs [9, 11,12,13,14,15,16,17, 20]

Summary

Background

TFIID/SAGA histone fold-containing subunits contain conserved additional domains of unknown function. TAF6 Conserved C-terminal Domain Is Formed of HEAT Repeats of TFIIA, TFIID binds to a TATA box-containing promoter by inducing the formation of a DNA loop This process appears to lock TFIID onto promoter DNA, suggesting the first steps of the assembly of the transcription preinitiation complex. This result is, in contradiction with the fact that a human TFIID complex incorporating the TAF6 isoform TAF6␦, which lacks the central part of its histone fold domain, still retains all TAFs except TAF9 [27], suggesting that other regions of TAF6 are required for integration of this TAF within TFIID To address this issue, we have performed biochemical and structural studies on the conserved C-terminal region of TAF6. Our results suggest that (i) formation of histone fold-containing heterodimers within TFIID does not rely on the histone fold motif of each partner and (ii) TFIID assembly is an intricate process possibly requiring conformational changes that may be important for TFIID function

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION