9.3

The mechanisms that regulate transcription initiation and elongation by RNA polymerase [blank_start]II[blank_end] have been studied extensively because this polymerase is the one that transcribes mRNAs.

Today the transcription start site for a newly characterized mRNA is generally determined simply by identifying the DNA sequence encoding the 3′-capped nucleotide of the encoded mRNA.

Several different types of DNA sequences can function as [blank_start]promoters[blank_end] for RNA polymerase II

Several different types of DNA sequences can function as promoters for RNA polymerase II, telling the polymerase where to initiate transcription of an RNA complementary to the template strand of a double-stranded DNA molecule. These sequences include [blank_start]TATA[blank_end] boxes, [blank_start]initiators[blank_end], and [blank_start]CpG[blank_end] islands.

In all these highly transcribed genes, a conserved sequence called the TATA box was found about 26–31 bp upstream of the transcription start site (Figure 9-16). Mutagenesis studies have shown that a singlebase change in this nucleotide sequence drastically [blank_start]decreases[blank_end] in vitro transcription of the gene adjacent to it.

If the base pairs between the TATA box and the normal transcription start site are deleted, transcription begins at the same point regardless.

In contrast to the conserved TATA box sequence, however, only an extremely degenerate initiator consensus sequence has been defined

CG-rich sequences are bound by histone octamers more [blank_start]weakly[blank_end] than CG-poor sequences because [blank_start]more[blank_end] energy is required to bend them into the small-diameter loops required to wrap around the histone octamer forming a [blank_start]nucleosome[blank_end] (see Figure 8-24). As a consequence, CpG islands coincide with nucleosome-[blank_start]free[blank_end] regions of DNA.

a current hypothesis is that the general transcription factors discussed in the next section can bind to them because CpG islands [blank_start]exclude[blank_end] nucleosomes.

Another remarkable feature of CpG islands is that transcription from these elements is initiated in both directions, even though only transcription of the sense strand yields an mRNA.

Initiation of transcription by RNA polymerase II requires several initiation factors.

Initiation Factors: They are called [blank_start]general transcriptionfactors[blank_end] because they are required at most, if not all, promoters of genes transcribed by RNA polymerase [blank_start]II[blank_end].

These factors that regulate elongation in the promoter-proximal region provide a mechanism for controlling gene transcription in addition to the regulation of transcription initiation. This overall strategy for regulating transcription at both the [blank_start]initiation[blank_end] and [blank_start]elongation[blank_end] steps in the promoter-proximal region is similar to the regulation of the trp operon in E. coli (see Figure 9-7), although the molecular mechanisms involved are distinct.

Three principal types of promoter sequences have been identified in eukaryotic DNA. The [blank_start]TATA box[blank_end] is prevalent in highly transcribed genes. [blank_start]Initiator[blank_end] promoters are found in some genes, and [blank_start]CpG islands[blank_end], the promoters for about 70 percent of protein-coding genes in vertebrates, are characteristic of genes transcribed at a [blank_start]low[blank_end] rate.

Transcription of protein-coding genes by Pol II is initiated by sequential binding of the following in the indicated order: [blank_start]TFIID[blank_end], which contains the TBP subunit that binds to TATA box DNA; [blank_start]TFIIA[blank_end] and [blank_start]TFIIB[blank_end]; a complex of Pol II and TFIIF; TFIIE; and finally, [blank_start]TFIIH[blank_end] (see Figure 9-19).

The helicase activity of a TFIIH subunit helps to separate the DNA strands at the transcription start site in most promoters, a process that requires hydrolysis of ATP. As Pol II begins transcribing away from the start site, its [blank_start]CTD[blank_end] is phosphorylated on [blank_start]serine 5[blank_end] by the [blank_start]TFIIH[blank_end] kinase domain.

In metazoans, [blank_start]NELF[blank_end] and [blank_start]DSIF[blank_end] associate with Pol II after initiation, inhibiting elongation fewer than 100 bp from the transcription start site. Inhibition of elongation is relieved when [blank_start]cyclin T–CDK9[blank_end] (also called P-TEFb) associates with the [blank_start]elongation complex[blank_end] and CDK9 [blank_start]phosphorylates[blank_end] subunits of NELF, DSIF, and serine 2 of the Pol II CTD.