Pol II Preinitiation Complex (Pol II and Company)

In order to initiate transcription, general transcription factors are required to place Pol II at the transcription start site, which is a promoter containing a TATA box. They also help separate DNA strands so that the template strand can enter the active site of the enzyme. As these initiation factors are necessary for most, if not all, promoters transcribed by RNA Polymerase II, they are called general transcription factors. Recall that transcription factors are proteins. Together, these transcription factors form what is referred to as a preinitiation complex in vivo and in vitro. We are going to explain the in vitro preinitiation complex and the steps of the order of various transcription factor binding.

1) The first transcription factor to bind to the TATA box, is not shockingly, the TATA box binding protein, or TBP. It folds into a saddle-like structure, because of its C-terminal domain, and from there, it interacts with the minor groove of DNA. It bends the helix significantly.

As the DNA-binding surface of the TBP is highly conserved in eukaryotes, it helps explain the high conservation of the TATA box promoter element.

2) Next, TFIIB binds by its C-terminal domain making contact with both TBP and either side of the TATA box. The N-terminal domain is inserted into the RNA exit channel of Pol II and helps Pol II in melting the DNA strands at the transcription start site. It also interacts with the template strand near the Pol II active site.

3) TFIIF (heterodimer) and Pol II bind, which positions Pol II over the start site.

4) After, that TFIIE (tetrameric with two different subunits) binds next to TFIIF and PolII on the side closest to the upstream direction. This creates a docking site for TFIIH.

5) Now, TFIIH (10 different subunits) binds, completing the assembly of the Pol II Preinitiation Complex.

6) Helicase activity of one of the TFIIH subunits uses energy from ATP hydrolysis to help unwind the DNA duplex at the start site, which allows Pol II to form an open complex (transcription bubble). There is a release of the general transcription factors, except for TBP. Pol II elongates the nascent mRNA strand, and has a phosphorylated CTD, which allows for association of the enzymes required to add the 5' cap.

Brief note of some differences in vivo:

In vivo, TBP is part of TFIID, which is a complex of not only TBP, but also 13 other subunits called TAFs. Also, TFIIA is required to form the preinitiation complex and binds to TBP and the TATA box.