FIGURE 12-12 Pol II core promoter The figure shows the positions of various DNA elements relative to tfie transcription start site (indicated by the arrow above the DNA) These elements, described in the text, are as follows: BRE (TRffi recognition element); TATA (IATA Bcix), Inr {initiator element), and DPE (downstream promoter element). Also shown (below) ate the consensus sequence for each element (determined in the same way as described for the bacterial promoter elements, see Bra 12 )), and (above) the name of the general transcription factor that recognize*, cadi element. (Source Butler J.E.P. et a!. 2002. Genes and Development 16: 2583-2592, Fig I.)

Beyond—and typically upstream of—the core promoter, there are other sequence elements required for efficient transcript)on in vivo. Together these elements constitute the regulatory sequences and can be grouped into various categories, reflecting their location, and the organism in question, as much as their Junction. These elements include: promoter proximal elements; upstream activator sequences (UASs); enhancers; and a series of repressing elements called silencers, boundary elements, end insutators. All these DNA elements bind regulatory proteins (activators and repressors), which help or hinder transcription from the core promoter, the subject of Chapter 17. Some of these regulatory sequences can be located many 10s or even 100s of Kb from the core promoters on which they act.

RNA Polymerase II Forms a Pre-lnitiation Complex with General Transcription Factors at the Promoter

The general transcription factors collectively perform I he functions performed by o in bacterial transcription, despite showing no significant sequence homology to that protein. Thus, the general transcription factors help polymerase bind to the promoter and melt the DNA (comparable to the transition from closed to open complex in the bacterial case). They also help polymerase escape from the promoter and embark on the elongation phase. The complete set of general transcription factors and polymerase, bound together at the promoter and poised for initiation, is called the pre-initiation complex.

As we described above (and in Figure 12-12) many Pol II promoters contain a so-called TATA element (some 30 base pairs upstream from the transcription start site). This is where pre-initiation complex formation begins. The TATA element is recognized by the general transcription factor called TFIID. (The nomenclature "TFll" denotes a transcription factor for Pol U. with individual factors distinguished as A, B, and so on.) Like many of the general transcription faclors, TFIID is in fact a multi-subunit complex. The component of TFIID that binds to the TATA DNA sequence is called TBP (TATA binding protein). The other subunits in this complex are called TAFs, For TBP associated factors. Some TAFs help bind the DNA at curiam promoters, and others control the DNA-binding activity of TBP.

Upon binding DNA, TBP extensively distorts the TATA sequence (we shall discuss this event in more detail presently). The resulting TBP-DNA complex provides a platform to recruit other genera!

transcription factors and polymerase itself to the promoter. In vitro, these proteins assemble at the promoter in the following order (Figure 12-13): TFIIA, TFI1B, TFIIF together With polymerase (in complex with yet more proteins, such as those in the Mediator Complex, which we describe below), and then TFIIE and TFII1I, which bind upstream of PoJ fl. Formation of the pre-initiation complex containing these components is followed by promoter melting. In contrast lo the situation in bacteria, promoter melting in eukaryoles requires hydrolysis of ATP and is mediated by TFIIH. It is the helicase-like activity of that factor which stimulates unwinding of promoter DNA,

FIGURE 12-13 Transcription initiation by RNA polymerase II. The step-wise assembly of the Pot II preinitiation complex ts shown here, and described in detail in the text Once assembled ai the promoter. Pol II leaves tlte pre initiation complex upon addition of the nucleotide precursors required for RWA synthesis, and after phosphorylation of Ser resides within Itie enzyme's tail." The tail contains multiple repeats of the heptapeptide sequence: Tyr-Set Pro-Tnr-Ser Pro-Ser {see Figure 12-18).

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