Promoter Organization

Transcription initiation rates - In E. coli, rates of transcription initiation vary enormously--from about one initiation every 10 seconds for some genes to as infrequently as once per generation (30 to 60 minutes) for others. Because all genes in bacteria are transcribed by the same core enzyme, variations in promoter structure must be largely responsible for the great variation in the frequency of initiation. Variations in promoter structure represent a simple way for the cell to vary rates of transcription from different genes.

Common sequence motifs - By analyzing DNA sequences ahead of genes, it is possible to identify common sequence features of promoter regions. For instance, near position -10 (position +1 is the start site of transcription), a common sequence motif is present in E. coli that is close to (or exactly) the sequence TATAAT on the sense strand (nontranscribed DNA strand). Another region of conserved nucleotide sequence is centered at nucleotide -35, with a consensus sequence of TTGACA. No known natural promoter has -35 and -10 regions that are identical to the consensus sequences, but in general, the more closely these regions in a promoter resemble the consensus sequences, the more efficient that promoter is in initiating transcription. Figure 26.12 indicates the extent to which different nucleotides are conserved.

Mutations and spacing - Mutations in promoter regions affect transcription efficiency in vivo. As shown in Figure 26.12, most of the promoter mutations that have been sequenced change the structure of either the -35 region or the -10 region, pointing directly to those sequences as having the greatest effect on transcriptional initiation efficiency. Although most natural promoters have a 17-nucleotide spacer between the -35 and -10 regions, many have 16 or 18. In vitro studies show that a 17-nucleotide spacer yields the most efficient promoter structure.

RNA polymerase contacts - The -35 region and the -10 region, plus a few nucleotides upstream of -10, are the major contact points for RNA polymerase in an open-promoter complex. The RNA polymerase subunit also makes contact in the -40 to -60 region. Figure 26.14 summarizes the results of these contacts for a promoter. Because there are two turns of the helix between the -35 and -10 regions, RNA polymerase is postulated to bind to DNA primarily on one side of the duplex. The data on nucleotide reactivity support this conclusion.

Super helicity - Another factor affecting transcriptional efficiency, in addition to the base sequence of the promoter, is the superhelical tension on the DNA template. The relation between DNA topology and transcriptional efficiency is not completely clear, because the transcription of some genes is activated in vivo when the template is highly supercoiled, but the transcription of other genes is inhibited under the same conditions. Interestingly, the promoter for transcription of DNA gyrase subunits becomes activated when the gene is in a relaxed state. Given that gyrase introduces superhelical turns, this finding seems to represent a feedback mechanism in which the cell responds appropriately to a signal that intracellular DNA is becoming too relaxed.

See also: Initiation and Elongation, Structure of RNA Polymerase, Transcription Regulation in Phage, Lactose Operon Regulation, Supercoiling (from Chapter 4)

INTERNET LINK: Supercoiling