Bacterial RNA polymerase is composed of a core of subunits (β, β′, α1, α2, ω), which have RNA synthesizing activity, and a specificity factor (σ), which identifies the start of transcription by recognizing and binding to sequence elements within promoter DNA. Four core promoter consensus sequences, the −10 element, the extended −10 (TGn) element, the −35 element, and the UP elements, have been known for many years; the importance of a nontemplate G at position −5 has been recognized more recently. However, the functions of these elements are not the same. The AT-rich UP elements, the −35 elements (−35TTGACA−30), and the extended −10 (−15TGn−13) are recognized as double-stranded binding elements, whereas the −5 nontemplate G is recognized in the context of single-stranded DNA at the transcription bubble. Furthermore, the −10 element (−12TATAAT−7) is recognized as both double-stranded DNA for the T:A bp at position −12 and as nontemplate, single-stranded DNA from positions −11 to −7. The single-stranded sequences at positions −11 to −7 as well as the −5 contribute to later steps in transcription initiation that involve isomerization of polymerase and separation of the promoter DNA around the transcription start site. Recent work has demonstrated that the double-stranded elements may be used in various combinations to yield an effective promoter. Thus, while some minimal number of contacts is required for promoter function, polymerase allows the elements to be mixed and matched. Interestingly, which particular elements are used does not appear to fundamentally alter the transcription bubble generated in the stable complex. In this review, we discuss the multiple steps involved in forming a transcriptionally competent polymerase/promoter complex, and we examine what is known about polymerase recognition of core promoter elements. We suggest that considering promoter elements according to their involvement in early (polymerase binding) or later (polymerase isomerization) steps in transcription initiation rather than simply from their match to conventional promoter consensus sequences is a more instructive form of promoter classification.

Summary The enterobactin system for iron transport in Escherichia coli is well characterized with the exception of the mechanism of enterobactin secretion to the extracellular environment. Escherichia coli membrane protein P43, encoded by ybdA in the chromosomal region of genes involved in enterobactin synthesis, shows strong homology to the 12‐transmembrane segment major facilitator superfamily of export pumps. A P43‐null mutation was created and siderophore nutrition assays, performed with a panel of E. coli strains expressing one or more outer membrane receptors for enterobactin‐related compounds, demonstrated that the P43 mutant was unable to secrete enterobactin efficiently. Products released from the mutant strain were identified with thin‐layer chromatography (TLC) and high‐performance liquid chromatography (HPLC), revealing that the P43 mutant secretes little, if any, enterobactin, but elevated levels of enterobactin breakdown products 2,3‐ dihydroxybenzoylserine (DHBS) monomer, dimer, and trimer. These data establish that P43 is a critical component of the E. coli enterobactin secretion machinery and provides a rationale for the designation of the previous genetic locus ybdA as entS to reflect its relevant biological function.

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Activation of bacteriophage T4 middle promoters, which occurs about 1 min after infection, uses two phage-encoded factors that change the promoter specificity of the host RNA polymerase. These phage factors, the MotA activator and the AsiA co-activator, interact with the s 70 specificity subunit of Escherichia coli RNA polymerase, which normally contacts the "10 and "35 regions of host promoter DNA. Like host promoters, T4 middle promoters have a good match to the canonical s 70 DNA element located in the "10 region. However, instead of the s 70 DNA recognition element in the promoter's "35 region, they have a 9 bp sequence (a MotA box) centred at "30, which is bound by MotA. Recent work has begun to provide information about the MotA/AsiA system at a detailed molecular level. Accumulated evidence suggests that the presence of MotA and AsiA reconfigures protein-DNA contacts in the upstream promoter sequences, without significantly affecting the contacts of s 70 with the "10 region. This type of activation, which is called 's appropriation', is fundamentally different from other well-characterized models of prokaryotic activation in which an activator frequently serves to force s 70 to contact a less than ideal "35 DNA element. This review summarizes the interactions of AsiA and MotA with s 70 , and discusses how these interactions accomplish the switch to T4 middle promoters by inhibiting the typical contacts of the C-terminal region of s 70 , region 4, with the host "35 DNA element and with other subunits of polymerase. OverviewUpon infection of Escherichia coli, bacteriophage T4 establishes its own developmental cycle. Within 20 min, the phage programmes the generation of approximately 200 copies of its genome, the packaging of that DNA, and finally its escape from the host by lysis (reviewed by Miller et al., 2003). Regulation of this cycle is achieved largely by phage promoters, which sequentially express early, middle and late phage genes. Because T4 does not encode its own RNA polymerase, it must direct the host transcriptional machinery to these phage promoters at the correct time during infection. T4 encodes factors that accomplish this takeover by altering the specificity of the host E. coli RNA polymerase as infection proceeds (reviewed by Miller et al., 2003;Stitt & Hinton, 1994).E. coli RNA polymerase consists of a core of five subunits (a 2 , b, b9 and v), which contains the RNA-synthesizing activity, and a s factor that binds to a specific promoter sequence and sets the start site for transcription (reviewed by Gruber & Gross, 2003;Paget & Helmann, 2003 and a 235 element, having a consensus sequence of 59-TTGACA-39 (Campbell et al., 2002;Gardella et al., 1989;Keener & Nomura, 1993;Murakami et al., 2002b;Siegele et al., 1989;Vassylyev et al., 2002;Waldburger et al., 1990).(All sequences are given as the top, i.e. the non-template, strand of DNA.) To a first approximation, the strength of a host promoter reflects the match between its 210 and 235 sequences and the canonical sequences for these region...