Transcription factor TFIIA

J. J. Kang, D. T. Auble, J. A. Ranish, and S. Hahn. Analysis of the yeast transcription factor TFIIA distinct functional regions and a polymerase n-spectfic role in basal and activated transcription. Mol Cell Bid, IS (3), 1234-1243, 1995. 

Ftgure 29.20 Transcription initation. Transcription factors TFIIA. B, D, E, F. and H are essential in initiating transcription by RNA polymerase II. The step-by-step assembly of these general transcription factors begins with the binding of TFIID (purple) to the TATA box. 

Fig. 9-4 Promoters interact with distant enhancers via iooping, faciiitated by the mediator compiex. (A) DNA iooping around so its enhancer sequences, via activator transcription factors, interact with the compiex that forms around RNA poiymerase ii. (B) RNA poiymerase ii, iike other eukaryotic RNA poiymerases, is recruited by transcription factors which assembie step-wise at the promoter, its C terminai taii is phosphoryiated to recruit eionga-tion factors which permit it to ieave the promoter and commence transcription. The various transcription factors (TFiiA, TFiiB, etc.) are indicated by their abbreviated names, uniike FiAT which is histone acetyi transferase.

TBP has been cloned from many organisms, ranging from archeabacteria 4, 5) to humans (see (6) for a review). Crystallographic determinations of TBP structures from archeabacteria (7), yeast (5), and plants (6) reveal the same architecture a molecular saddle with a near two-fold symmetry axis. TBP has also been crystallized in complex with DNA (9-72), and in ternary complexes with two other basic transcription factors, TFIIA (75, 14) and TFIIB (75). In all the crystallized complexes, TBP binds to 8 basepairs in the minor groove of the DNA encoding a characteristic TA repeat. The underside of the TBP saddle is the binding interface with the DNA. This interface is mostly hydrophobic, but 6 hydrogen bonds have been identified in the crystal structures. 

The DNA part of each control module can be divided into three main regions, the core or basal promoter elements, the promoter proximal elements and the distal enhancer elements (Figure 9.1). The best characterized core promoter element is the TATA box, a DNA sequence that is rich in A-T base pairs and located 25 base pairs upstream of the transcription start site. The TATA box is recognized by one of the basal transcription factors, the TATA box-binding protein, TBP, which is part of a multisubunit complex called TFIID. This complex in combination with RNA polymerase 11 and other basal transcription factors such as TFIIA and TFIIB form a preinitiation complex for transcription. 

TFIIA and TFIIB are two basal transcription factors that are involved in the nucleation stages of the preinitiation complex by binding to the TBP-TATA box complex. Crystal structures of the ternary complex TFIIA-TBP-TATA box have been determined by the groups of Paul Sigler, Yale University, and Timothy Richmond, ETH, Zurich, and that of the TFIIB-TBP-TATA box by Stephen Burley and collaborators. The TBP-DNA interactions and the distortions of the DNA structure are essentially the same in these ternary complexes as in the binary TBP-TATA complex. 

Sequences farther upstream from the start site determine how frequently the transcription event occurs. Mutations in these regions reduce the frequency of transcriptional starts tenfold to twentyfold. Typical of these DNA elements are the GC and CAAT boxes, so named because of the DNA sequences involved. As illustrated in Figure 37—7, each of these boxes binds a protein, Spl in the case of the GC box and CTF (or C/EPB,NF1,NFY) by the CAAT box both bind through their distinct DNA binding domains (DBDs). The frequency of transcription initiation is a consequence of these protein-DNA interactions and complex interactions between particular domains of the transcription factors (distinct from the DBD domains—so-called activation domains ADs) of these proteins and the rest of the transcription machinery (RNA polymerase II and the basal factors TFIIA, B, D, E, F). (See... 

RNA polymerases interact with unique cw-active regions of genes, termed promoters, in order to form preinitiation complexes (PICs) capable of initiation. In eukaryotes the process of PIC formation is facilitated by multiple general transcription factors (GTFs), TFIIA, B, D, E, F, and H. 

TATA-Binding Protein The first component to bind in the assembly of a preinitiation complex at the TATA box of a typical Pol II promoter is the TATA-binding protein (TBP). The complete complex includes the basal (or general) transcription factors TFIIB, TFIIE, TFIIF, TFIIH Pol II and perhaps TFIIA (not all of the factors are shown in Fig. 28-27). This minimal preinitiation complex, however, is often insufficient for the initiation of transcription and generally does not form at all if the promoter is obscured within chromatin. Positive regulation leading to transcription is imposed by the transactivators and coactivators. 

Phil Sharp and Leonard Guarente showed that at least four transcription factors are required in addition to polymerase II for initiation from the major late promoter of adenovirus. In vitro studies indicate that these factors assemble in an orderly fashion (see fig. 28.12b). First the TFIID complex binds to the TATA box. Sequential binding of TFIIA, TFIIB, RNA polymerase II, and TFIIE follow. It is believed that this multifactor complex functions for a large number of eukaryotic promoters that contain TATA boxes. 

Finally, in Plate 21, the structure of the yeast TFIIA-TBP-DNA complex is shown. TFIIA interacts with TBP and the TFIID complex and stimulates transcription of the Pol II gene.29-32 TFIIA has two characteristic structural motife. One is a six-stranded (i-sand-wich, the other is a left-handed four-helix bundle. The P-sandwich domain of TFUA is alone responsible for all of the interactions with the DNA, whereas its helix-bundle domain projects away and is free to interact with signal-responsive transcription factors. These interactions are important for regulation of transcription. Little conformational change occurs in TBP when it binds to TFIIA. The main difference between the TFIIA-TPB complex and the TFIIB-TBP complex is that TFIIA binds upstream of the TATA box, away from the transcriptional start site, whereas TFIIB binds downstream of the TATA box. Moreover, TFIIB is positioned on the side opposite to TFIIA. 

We have learnt from the structures that the TBP/TATA complex is a nucleoprotein scaffold upon which other factors, such as TFIIA and TFIIB attach with high affinity through a combination of stereospecific and electrostatic interactions. TFIIB is positioned at the transcriptional start site, between the attachment sites for TBP and Pol II. On the other hand, TFIIA has no contacts with the DNA downstream of the TATA box and does not interact with the transcriptional start site and/or any of the components of the basal transcription machinery, all of which are located downstream of the TATA box. Thus, TFIIA and TFIIB can bind simultaneously, without mutual interference. Moreover, TFIIA is accessible to specific, signal-responsive regulatory transcription factors. Its location upstream of the TATA box also enables TFIIA to absorb and scavenge transcriptional inhibitors, making them ineffectual. 

Cis-acting elements constitute only part of the puzzle of eukaryotic gene expression. Transcription factors that bind to these elements also are required. For example, RNA polymerase II is guided to the start site by a set of transcription factors known collectively as TFII (TF stands for transcription factor, and II refers to RNA polymerase II). Individual TFII factors are called TFIIA, TFIIB, and so on. Initiation begins with the binding of TFIID to the TATA box (Figure 28.19). 

TBP bound to the TATA box is the heart of the initiation complex (see Figure 28.19). The surface of the TBP saddle provides docking sites for the binding of other components (Figure 28.21). Additional transcription factors assemble on this nucleus in a defined sequence. TFIIA is recruited, followed by TFIIB and then TFIIF—an ATP-dependent helicase that initially separates the DNA duplex for the polymerase. Finally, RNA polymerase II and then TFIIE join the other factors to form a complex called the basal transcription apparatus. Sometime in the formation of this complex, the carboxyl-terminal domain of the polymerase is phosphorylated on the serine and threonine residues, a process required for successful initiation. The importance of the carboxyl-terminal domain is highlighted by the finding that yeast containing mutant polymerase II with fewer than 10 repeats is not viable. Most of the factors are released before the polymerase leaves the promoter and can then participate in another round of initiation. 

Recently, a family of related proteins has been identified in metazoa, the TBP-like proteins or TLPs [16], which are also able to bind to DNA, albeit not to the consensus TATA boxes. The physiological role of TLPs is an area of intense research, and some proposals suggest that it sequesters other general transcription factors (such as TFIIA), thereby repressing RNApol-II transcription mediated by TBP [17]. 

Ternary complexes formed by TBP, DNA and TFIIA or TFIIB have also been crystallized [27-32] and their structures are available in the PDB and NDB (see Table 1). These complexes display practically the same mode of interaction between TBP and the DNA moiety as found in the corresponding binary complexes. They also explain the inability of TFIIB to bind to DNA on its own, as it is found to contact the DNA upstream and downstream of TBP, an impossible feat in a straight DNA molecule. This feature places TBP among the architectural transcription factors, together with DBF, HMG, SRY and LEFl... 

How do the special transcription factors regulate the activities of the basal transcription factors Information on this issue is only beginning to become available. One might expect, for example, to find a scenario where RXR/VDR directly contacts TFIIA, to stimulate the activity of the initiation complex. 

A number of the core transcription factors are released and RNA polymerase II, together with TFIIF, moves along the DNA. A residual complex, containing the TATA binding protein (TBP), TFIIA, TATA binding associated factors (TAFs) and probably activator proteins, remains at the start site, ready to initiate another round of transcription. 

TFIID (which contains the TATA-box binding protein, TBP) binds to the TATA box. TFIIA and TFIIB then bind, followed by recruitment of RNA polymerase II and TFIIF. TFIIH and TFIIE then bind to form the preinitiation complex (PIC). Kinases phosphorylate the C-terminal domain of Pol II, leading to the open complex in which the DNA strands are separated. RNA is produced during elongation as Pol II and TFIIF leave the promoter and the other general transcription factors behind. Pol II dissociates during the termination phase, and the CTD is dephosphorylated. Pol II/TFIIF is then recycled to bind to another promoter. 

TFIIA, TFIIB, TFIID, TFIIE, TFIIF, and TFIIH are the general transcription factors. TFIID is also the TATA-box binding protein and is associated with TAFs (TBP associated factors). 

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