Transcription factors

The specific arrangement of two a helices joined by a loop region in lambda Cro and repressor, as well as in CAP, constitute the helix-turn-helix DNA-binding motif (Figure 8.8), which also occurs in some eucaryotic transcription factors as discussed in Chapter 9. The orientation of the two helices and... 

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. 

The promoter proximal elements are usually 100 to 200 base pairs long and relatively close to the site of initiation of transcription. Within each of these elements there are DNA sequences specifically recognized by several different transcription factors which either interact directly with the preinitiation complex or indirectly through other proteins. 

Figure 9.2 Schematic model for transcriptional activation. The TATA box-binding protein, which bends the DNA upon binding to the TATA box, binds to RNA polymerase and a number of associated proteins to form the preinitiation complex. This complex interacts with different specific transcription factors that bind to promoter proximal elements and enhancer elements.

The general transcription factor TFllD is believed to be the key link between specific transcription factors and the general preinitiation complex. However, the purification and molecular characterization of TFllD from higher eucaryotes have been hampered by its instability and heterogeneity. All preparations of TFllD contain the TATA box-binding protein in combination with a variety of different proteins called TBP-associated factors, TAFs. When the preinitiation complex has been assembled, strand separation of the DNA duplex occurs at the transcription start site, and RNA polymerase II is released from the promoter to initiate transcription. However, TFIID can remain bound to the core promoter and support rapid reinitiation of transcription by recruiting another molecule of RNA polymerase. 

The polypeptide chains of the specific transcription factors usually have two different functions one is to bind to a specific DNA sequence and another is to activate transcription. These two functions are often... 

The two homologous repeats, each of 88 amino acids, at both ends of the TBP DNA-binding domain form two stmcturally very similar motifs. The two motifs each comprise an antiparallel p sheet of five strands and two helices (Figure 9.4). These two motifs are joined together by a short loop to make a 10-stranded p sheet which forms a saddle-shaped molecule. The loops that connect p strands 2 and 3 of each motif can be visualized as the stirmps of this molecular saddle. The underside of the saddle forms a concave surface built up by the central eight strands of the p sheet (see Figure 9.4a). Side chains from this side of the P sheet, as well as residues from the stirrups, form the DNA-binding site. No a helices are involved in the interaction area, in contrast to the situation in most other eucaryotic transcription factors (see below). 

The side of the p sheet that faces away from DNA is covered by two long a helices. One of these helices contains a number of basic residues from the middle segment of the polypeptide chain while the second helix is formed by the C-terminal residues. Residues from these two helices and from the short loop that joins the two motifs (red in Figure 9.4) are likely candidates for interactions with other subunits of the TFIID complex, and with specific transcription factors. 

The sharp bend of DNA at the TATA box induced by TBP binding is favorable for the formation of the complete DNA control module in particular, for the interaction of specific transcription factors with TFIID. Since these factors may bind to DNA several hundred base pairs away from the TATA box, and at the same time may interact with TBP through one or several TAFs, there must be several protein-DNA interactions within this module that distort the regular B-DNA structure (see Figure 9.2). The DNA bend caused by the binding of TBP to the TATA box is one important step to bring activators near to the site of action of RNA polymerase. 

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. 

Eucaryotes have many more genes and a broader range of specific transcription factors than procaryotes and gene expression is regulated by using sets of these factors in a combinatorial way. Eucaryotes have found several different solutions to the problem of producing a three-dimensional scaffold that allows a protein to interact specifically with DNA. In the next chapter we shall discuss some of the solutions that have no counterpart in procaryotes. However, the procaryotic helix-turn-helix solution to this problem (see Chapter 8) is also exploited in eucaryotes, in homeodomain proteins and some other families of transcription factors. 

Homeoboxes code for homeodomains, sequences of 60 amino acids that function as the DNA-binding regions of transcription factors. Each homeo-box gene in Drosophila is expressed only in its own characteristic subset of embryonic cells, and almost every embryonic cell contains a unique combination of homeodomain proteins. 

In vivo specificity of homeodomain transcription factors depends on interactions with other proteins... 

Figure 9.12 Schematic diagram of the structure of the heterodimeric yeast transcription factor Mat a2-Mat al bound to DNA. Both Mat o2 and Mat al are homeodomains containing the helix-turn-helix motif. The first helix in this motif is colored blue and the second, the recognition helix, is red. (a) The assumed structure of the Mat al homeodomain in the absence of DNA, based on Its sequence similarity to other homeodomains of known structure, (b) The structure of the Mat o2 homeodomain. The C-terminal tail (dotted) is flexible in the monomer and has no defined structure, (c) The structure of the Mat a 1-Mat a2-DNA complex. The C-terminal domain of Mat a2 (yellow) folds into an a helix (4) in the complex and interacts with the first two helices of Mat a2, to form a heterodimer that binds to DNA. (Adapted from B.J. Andrews and M.S. Donoviel, Science 270 251-253, 1995.)...

The polypeptide chain of p53 is divided in three domains, each with its own function (Figure 9.16). Like many other transcription factors, p53 has an N-terminal activation domain followed by a DNA-binding domain, while the C-terminal 100 residues form an oligomerization domain involved in the formation of the p53 tetramers. Mutants lacking the C-terminal domain do not form tetramers, but the monomeric mutant molecules retain their sequence-specific DNA-binding properties in vitro. 

DNA binding by TBP is strongly dependent on the presence of T-A base pairs in the TATA box. Bending allows remote sites on the DNA, with their bound cognate specific transcription factors, to come close together such that the proteins can interact to form the transcriptional preinitiation complex. 

Burley, S.K., Boeder, R.G. Biochemistry and structural biology of transcription factor IID. Anna. Rev. Bioehem. 

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