RNA Pol II Genes: Promoters And Enhancers
▶RNA polymerase II: RNA polymerase II (RNA Pol II) is located in the nucleoplasm. It is responsible for the transcription of all protein-coding genes, some small nuclear RNA genes and sequences encoding micro RNAs and short interfering RNAs. The pre mRNAs must be processed after synthesis by cap formation at the 5’-end of the RNA and poly(A) addition at the 3_-end, as well as removal of introns by splicing .
▶Promoters: Many eukaryotic promoters contain a sequence called the TATA box around 25–35 bp upstream from the start site of transcription. It has the 7 bp consensus sequence 5’-TATA(A/T)A(A/T)-3’ although it is now known that the protein which binds to the TATA box, TBP, binds to an 8 bp sequence that includes an additional downstream base pair, whose identity is not important.
The TATA box acts in a similar way to an E. coli promoter –10 sequence to position the RNA Pol II for correct transcription initiation. While the sequence around the TATA box is critical, the sequence between the TATA box and the transcription start site is not critical. However, the spacing between the TATA box and the start site is important. Around 50% of the time, the start site of transcription is an adenine residue. Some eukaryotic genes contain an initiator element instead of a TATA box.
The initiator element is located around the transcription start site. Many initiator elements have a C at position –1 and an A at +1. Other promoters have neither a TATA box nor an initiator element. These genes are generally transcribed at low rates, and initiation of transcription may occur at different start sites over a length of up to 200 bp. These genes often contain a GC-rich 20–50 bp region within the first 100–200 bp upstream from the start site.
▶Upstream regulatory elements: The low activity of basal promoters is greatly increased by the presence of other elements located upstream of the promoter. These elements are found in many genes which vary widely in their levels of expression in different tissues. Two common examples are the SP1 box, which is found upstream of many genes both with and without TATA boxes, and the CCAAT box. Promoters may have one, both or multiple copies of these sequences. These sequences which are often located within 100–200 bp upstream from the promoter are referred to as upstream regulatory elements (UREs) and play an important role in ensuring efficient transcription from the promoter.
▶Enhancers: Transcription from many eukaryotic promoters can be stimulated by control elements that are located many thousands of base pairs away from the transcription start site. This was first observed in the genome of the DNA virus SV40. A sequence of around 100 bp from SV40 DNA can significantly increase transcription from a basal promoter even when it is placed far upstream. Enhancer sequences are characteristically 100–200 bp long and contain multiple sequence elements which contribute to the total activity of the enhancer. They may be ubiquitous or cell type-specific. Classically, enhancers have the following general characteristics:
- they exert strong activation of transcription of a linked gene from the correct start site.
- They activate transcription when placed in either orientation with respect to linked genes.
- They are able to function over long distances of more than 1 kb whether from an upstream or downstream position relative to the start site.
- They exert preferential stimulation of the closest of two tandem promoters.
However, as more enhancers and promoters have been identified, it has been shown that the upstream promoter and enhancer motifs overlap physically and functionally. There seems to be a continuum between classic enhancer elements and those promoter elements which are orientation specific and must be placed close to the promoter to have an effect on transcriptional activity.
▶RNA polymerase II: RNA polymerase II (RNA Pol II) catalyzes the synthesis of the mRNA precursors for all protein-coding genes. RNA Pol II-transcribed pre-mRNAs are processed through cap addition, poly(A) tail addition and splicing.
▶Promoters: Many RNA Pol II promoters contain a sequence called a TATA box which is situated 25–30 bp upstream from the start site. Other genes contain an initiator element which overlaps the start site. These elements are required for basal transcription complex formation and transcription initiation.
▶Upstream regulatory elements: Elements within the 100–200 bp upstream from the promoter are generally required for efficient transcription. Examples include the SP1 and CCAAT boxes.
▶Enhancers: These are sequence elements which can activate transcription from thousands of base pairs upstream or downstream. They may be tissue-specific or ubiquitous in their activity and contain a variety of sequence motifs. There is a continuous spectrum of regulatory sequence elements which span from the extreme long-range enhancer elements to the short-range promoter elements.