RNA polymerase alpha subunit binding site in positively controlled promoters: a new model for RNA polymerase-promoter interaction and transcriptional activation in the Escherichia coli ada and aidB genes.

Landini, Paolo; Volkert, Michael R.

doi:10.1002/j.1460-2075.1995.tb00107.x

Cited by 45 publications

(49 citation statements)

References 27 publications

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“…Although RNAP is bound to the malT promoter, CRP might be required to accelerate the formation of the open complex (43). Similarly, Ada does not enhance RNAP binding to the ada and aidB promoters but instead stimulates transcription by stabilizing an intermediate RNAP-promoter complex (44). Involvement of the ␣-CTD in the transition from closed to intermediate complexes was also proposed as the role of the p4 protein in activation at the phage phi29 A3 promoter (45).…”

Section: Discussionmentioning

confidence: 99%

A regulatory protein that interferes with activator-stimulated transcription in bacteria

Nakano

Zhang

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

129

218

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Transcriptional activator proteins in bacteria often operate by interaction with the C-terminal domain of the ␣-subunit of RNA polymerase (RNAP). Here we report the discovery of an ''anti-␣'' factor Spx in Bacillus subtilis that blocks transcriptional activation by binding to the ␣-C-terminal domain, thereby interfering with the capacity of RNAP to respond to certain activator proteins. Spx disrupts complex formation between the activator proteins ResD and ComA and promoter-bound RNAP, and it does so by direct interaction with the ␣-subunit. ResD-and ComA-stimulated transcription requires the proteolytic elimination of Spx by the ATPdependent protease ClpXP. Spx represents a class of transcriptional regulators that inhibit activator-stimulated transcription by interaction with ␣.Spx ͉ RNA polymerase ͉ ␣-subunit ͉ Bacillus subtilis ͉ transcriptional activation T ranscriptional activation in bacteria involves contacts between DNA-bound activators and promoter-bound RNA polymerase (RNAP). Most such interactions require the ␣-subunit of RNAP, which possesses several activator-interaction surfaces within its C-terminal domain (CTD) (1, 2). One such activator is ComA of the bacterium Bacillus subtilis (3-5), a response regulator, required for transcription of genes involved in the development of genetic competence (6). In response to high cell density, ComA becomes phosphorylated by interaction with its cognate histidine kinase, ComP, that is activated when it binds the pheromone ComX (7-9). ComA then activates the transcription initiation of the srf operon, which encodes the competence regulatory peptide ComS (10, 11). ComS serves to release the transcriptional activator ComK from its inhibitory complex composed of the proteins MecA and ClpCP (10-13), so that ComK can stimulate transcription of genes required for DNA uptake in competent cells (14). Interestingly, ComAdependent transcription of srf requires the ATP-dependent protease ClpXP (15, 16), which functions to eliminate the 15.4-kDa Spx protein (refs. 17 and 18; see Results). A mutation in clpX blocks ComA-activated transcription and has severe effects on growth and development (17). These pleiotropic effects of ClpXP absence can be suppressed either by the elimination of Spx or by missense mutations in the rpoA gene that encodes the RNAP ␣-subunit (refs. 16 and 17; see Results). This latter finding suggested that Spx exerts its negative effect on ComA-mediated transcription, and other transcriptional activation systems, by interaction with RNAP. A similar relationship between Spx and the ␣-subunit of RNAP was observed for ResD-activated transcription (see Results). ResD, like ComA, is a response regulator and transcriptional activator. It is part of the ResDE two-component signal transduction system that is required for the transcription of genes that are induced in response to oxygen limitation (19).In this article we show that Spx interferes with activatorstimulated transcription by interaction with the RNAP ␣-CTD, a mechanism of transcriptional repression ...

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Section: Discussionmentioning

confidence: 99%

A regulatory protein that interferes with activator-stimulated transcription in bacteria

Nakano

Zhang

et al. 2003

Proc. Natl. Acad. Sci. U.S.A.

129

218

View full text Add to dashboard Cite

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“…The DNA promoter region shows characteristics of a promoter UP element, which are constituted by AT-rich regions of 20 bp spanning from ¹40 to ¹60 with a possible non-perfect inverted repeat. It has been demonstrated that these UP elements are the target sites of the C-terminal domain ᮊ 1997 Blackwell Science Ltd, Molecular Microbiology, 26, 361-372 of the ␣-subunit of RNA polymerase, thereby enhancing transcription (Ross et al, 1993;Attey et al, 1994;Landini and Volkert, 1995). In the case of the rrnB P 1 promoter of E. coli, interaction of ␣ with the UP element enhances promoter activity by approximately 30-fold (Ross et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Transcriptional analysis of the divergent cagAB genes encoded by the pathogenicity island of Helicobacter pylori

Spohn

Beier

Rappuoli

et al. 1997

Molecular Microbiology

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SummaryHelicobacter pylori strains isolated from most patients with peptic ulcer disease and adenocarcinoma express the vacuolating toxin VacA and contain a pathogenicity island named cag. The cag pathogenicity island codes for more than 40 putative proteins with features similar to bacterial secretion systems. One of these proteins, CagA, is an immunodominant antigen with unknown function encoded by the cagA gene. In the present study, we have analysed the functional promoter elements of the H. pylori cagA gene as well as of the divergently transcribed cagB gene. Primer extension analyses identified a single 5Ј end of the cagA mRNA, while two initiation sites were mapped in the case of the cagB mRNA. The promoters deduced upstream of these start points of transcription contained conserved ¹10 regions but no ¹35 regions with respect to the Escherichia coli 70 consensus sequence. Nevertheless, they could be activated in E. coli and in vitro by purified E. coli RNA polymerase. Deletion analyses indicated that the cagA and cagB genes are transcribed by overlapping promoters and that full activation requires sequences up to ¹70 and ¹96 respectively. Instead, basal transcription is likely to be mediated by ¹10 extended promoter-like sequences. RNA polymerase is able to bind the ¹40 to ¹60 region of the cagA promoter, and its binding is mediated by the ␣-subunit. This region resembles the UP elements of prokaryotic promoters in location, sequence and mechanism of interaction with the RNA polymerase. We discuss the features of these promoters and propose that they could represent a class of minimum promoters, which ensures a basic level of transcription, while full activation requires regulatory elements or a defined promoter context.

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“…At this promoter, truncation of RNA polymerase ␣CTD, or substitution of its R265 residue by alanine, abolishes ␣ binding to the UP element (14). These mutations also affect RNA polymerase binding to the Ϫ60 to Ϫ40 region of ada and aidB and impair transcription initiation (26). Based on these observations, the Ϫ60 to Ϫ40 regions of ada and aidB are functionally similar to the rrnB UP element and they are sufficient to recruit RNA polymerase.…”

Section: Mechanism Of Transcription Activation By Me Adamentioning

confidence: 97%

“…The evidence for this model was based on the observation that truncations in ␣CTD abolish transcription from the ada promoter in the presence of me Ada. However, more recent data indicate that RNA polymerase binds to the Ϫ60 to Ϫ40 region of the ada and aidB promoters via its ␣CTD, regardless of the presence of Ada (26). This region serves as the me Ada binding site, but it also closely resembles, in AϩT content, location, and function, the UP element transcription enhancer sequence identified in the rrnBP1 promoter (49).…”

Section: Mechanism Of Transcription Activation By Me Adamentioning

confidence: 99%

“…Based on these observations, the Ϫ60 to Ϫ40 regions of ada and aidB are functionally similar to the rrnB UP element and they are sufficient to recruit RNA polymerase. However, only upon binding of me Ada does the formation of a ternary complex that is proficient in transcription initiation take place (26). Thus, substitutions in ␣CTD result in reduced transcription from ada-dependent promoters because they prevent ␣ from binding the promoter Ϫ60 to Ϫ40 transcription enhancer sequence, rather than impairing its direct interaction with At the ada and aidB promoters, Ada interacts with a negatively charged patch in 70 : with the single exception of residue I590, substitutions affecting Ada-dependent transcription are in negatively charged amino acids (E574, E575, E591, E605, and D612).…”

Section: Mechanism Of Transcription Activation By Me Adamentioning

confidence: 99%

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Regulatory Responses of the Adaptive Response to Alkylation Damage: a Simple Regulon with Complex Regulatory Features

Landini

Volkert

2000

J Bacteriol

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2Alkylation damage to DNA occurs when cells encounter alkylating agents in the environment or when cellular metabolism produces active alkylators. To cope with DNA alkylation, cells have evolved genes that encode proteins with alkylationspecific DNA repair activities. In Escherichia coli, the main response specific for alkylation damage has been called the adaptive response (53). The adaptive response genes are induced upon exposure to exogenous alkylators by Ada-dependent induction, and also during stationary phase by rpoS-dependent gene expression, possibly to prevent accumulation of DNA damage due to increased endogenous production of alkylating agents. Recent studies of the regulatory mechanisms of Ada protein and the various responses of the individual promoters regulated by this protein has revealed a complexity of regulation not initially recognized. In this review we describe the roles of the Ada-regulated genes and the regulatory mechanisms that activate gene expression from the three Ada-dependent promoters. We will focus on Ada-dependent induction of the adaptive response genes, fine tuning of individual gene expression according to the growth phase, and the role played by Ada in shutting off the adaptive response. Ada-DEPENDENT REGULATION OF THE ADAPTIVE RESPONSE GENESThe adaptive response set of genes is comprised of the ada, alkA, alkB, and aidB genes. Expression of these genes is regulated by Ada, and their induction provides protection against alkylation damage to DNA. The ada gene product has both repair and regulatory activities. These two activities are closely tied to one another, as the Ada protein must be activated to perform its regulatory function and activation is a consequence of its DNA repair activity. Ada has two active methyl acceptor cysteine residues, Cys-69 and Cys-321, that are required for demethylation of DNA. Both sites can become methylated when Ada protein transfers the methyl group from the appropriate substrate DNA lesions to itself. This reaction is irreversible, and methylated Ada ( me Ada) is the terminal end product of the demethylation reaction (31). The two methyl acceptor sites present in Ada differ with respect to the lesions repaired. Cys-321 is the methyl acceptor site required for the removal of methyl groups from either O 6 -methylguanine or O 4 -methylthymine, two highly mutagenic lesions (10, 11). Cys-69 is required for demethylation of phosphomethyltriesters in the sugarphosphate backbone. This lesion is apparently innocuous, since Ada repairs only one of two stereoisomers (16,36,37,72), leaving the other to remain in DNA with no apparent deleterious consequences (28, 40). Although methylated phosphates are innocuous, this lesion is readily produced by methylating agents (37) and provides a sensitive regulatory signal that leads to induction of the Ada regulon. Once Ada protein transfers a methyl group from the methyl-phosphate to the Cys-69 residue, it becomes a transcriptional activator. Thus, the methylated phosphates in DNA serve as the signal that conv...

show abstract

RNA polymerase alpha subunit binding site in positively controlled promoters: a new model for RNA polymerase-promoter interaction and transcriptional activation in the Escherichia coli ada and aidB genes.

Cited by 45 publications

References 27 publications

A regulatory protein that interferes with activator-stimulated transcription in bacteria

A regulatory protein that interferes with activator-stimulated transcription in bacteria

Transcriptional analysis of the divergent cagAB genes encoded by the pathogenicity island of Helicobacter pylori

Regulatory Responses of the Adaptive Response to Alkylation Damage: a Simple Regulon with Complex Regulatory Features

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