Advances in bacterial promoter recognition and its control by factors that do not bind DNA

Abstract

Early work identified two promoter regions, the -10 and -35 elements, that interact sequence specifically with bacterial RNA polymerase (RNAP). However, we now know that several additional promoter elements contact RNAP and influence transcription initiation. Furthermore, our picture of promoter control has evolved beyond one in which regulation results solely from activators and repressors that bind to DNA sequences near the RNAP binding site: many important transcription factors bind directly to RNAP without binding to DNA. These factors can target promoters by affecting specific kinetic steps on the pathway to open complex formation, thereby regulating RNA output from specific promoters.

Figure 1. DNA elements and RNAP modules that contribute to promoter recognition by Eσ

The optimal UP element (α binding site) is an alternating series of A and T tracts (−57 to −38; 5′-AAAWWTWTTTTNNNAAANNN-3′; W = A or T; and N = any base). −35 refers to the −35 element, consensus sequence 5′-TTGACA-3′ from −35 to −30 (REF. 1). Ext refers to the extended −10 element, consensus sequence 5′-TGTG-3′ from −17 to −14 (REF. 50). −10 refers to the −10 element, consensus sequence 5′-TATAAT-3′ from −12 to −7 (REFS 1,2). Dis refers to the discriminator region, optimal sequence 5′-GGG-3′ from −6 to −4 (REFS 6,32,68). +1 is the transcription start site. The segments of the σ and α subunits of RNA polymerase (RNAP) that interact with these elements are described in the main text and REFS ,,,. αCTD, α carboxy-terminal domain.

Figure 2. Models of the open complex

a, b | Promoter elements and the regions of Eσ RNA polymerase (RNAP) that recognize promoter elements, the secondary channel and some other features of the enzyme are labelled. Segments of RNAP that bind to promoters are shown in space-fill, and ribbon representations of the DNA sections that are bound by these parts of RNAP are represented by matching colours (β, blue; β′, pink; α amino-terminal domain (αNTD), light grey; and ω, dark grey. The structures are adapted from the model of the open complex, which is based on the crystal structure of the fork–junction complex. The paths of the individual strands downstream from the −10 hexamer are based on modelling and biochemical studies^,, as DNA downstream of the −10 element is not present in the fork–junction complex. c | α carboxy-terminal domains (αCTDs) and upstream DNA are included. The αCTD structure is from REF. . The positions of the αCTDs and the path of the DNA upstream of the −35 hexamer are based on modelling and biochemical studies^,. Region 1.1 of σ was not resolved in the structure, but would be outside the main channel in the open complex. NTS, non-transcribed strand; TS, transcribed strand. Coordinates for the model from REF. were courtesy of R. Ebright and C. Lawson, Rutgers University, New Jersey, USA.

Figure 3. Steps in transcription initiation

In a, RP_C refers to the earliest promoter complex with RNA polymerase (RNAP), although this complex is not always kinetically detectable at all promoters. RP_O refers to the final complex before nucleoside 5′-triphosphate occupancy of RNAP. RP_I is used as an abbreviation for all the intermediates between RP_C and RP_O. The structures of these intermediates remain to be defined and could even differ at different promoters. k_a is the composite association-rate constant for RP_O formation. k_d is the composite dissociation constant. Individual steps are shown in b–f (REFS 15,30), and are described in more detail in the main text. The template DNA strand is in dark green; the non-template DNA strand is in light green; and the RNA transcript that emerges from the RNA channel is in red. The catalytic Mg²⁺ ion in the active site is indicated by a yellow sphere. Red circles represent acidic residues in regions 1.1 and 3.2, which are mobile modules in σ. In RP_C (b), the DNA is double stranded and downstream DNA has not moved into the main channel. In RP_I (c), σ region 1.1 begins to move out of, and downstream DNA begins to move into, the main channel. This probably occurs concurrent with DNA-strand separation^,, but the intermediates (RP_I) remain to be determined. In RP_O (d), the DNA strands have separated, with the template strand moving into position for base pairing with the first nucleoside 5′-triphosphate (NTP). In the ‘scrunched complex’ (REFS 34,35) (e), RNA synthesis has begun, and the DNA strands in or downstream of the −10 hexamer are temporarily ‘extruded’ out of their respective channels before promoter clearance. An NTP is shown entering the enzyme through the secondary channel. In the transcription elongation complex (TEC) (f), σ has dissociated from core RNAP. Figure adapted, with permission, from REF. (2003) Elsevier Science.