Helical phase dependent action of CRP: effect of the distance between the CRP site and the −35 region on promoter activity

Ushida, Chisato; Aiba, Hirofumi

doi:10.1093/nar/18.21.6325

Cited by 141 publications

(132 citation statements)

References 51 publications

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“…The linker connecting the N-and C-terminal domains of ␣ is sufficiently flexible to allow the CTD to move freely at least 30 bp along the promoter DNA. This is supported by previous reports that a single CRP site can be shifted from Ϫ61.5 to around Ϫ90, with retention of transcription activation (25) and with retention of the same critical determinants of CRP (26). NMR studies also indicate the presence of a flexible linker of at least 13 residues between the N-and C-terminal domains (27).…”

Section: Resultssupporting

confidence: 85%

Positioning of two alpha subunit carboxy-terminal domains of RNA polymerase at promoters by two transcription factors

Murakami

Owens

Belyaeva

et al. 1997

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

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Interactions between the cAMP receptor protein (CRP) and the carboxy-terminal regulatory domain (CTD) of Escherichia coli RNA polymerase ␣ subunit were analyzed at promoters carrying tandem DNA sites for CRP binding using a chemical nuclease covalently attached to ␣. Each CRP dimer was found to direct the positioning of one of the two ␣ subunit CTDs. Thus, the function of RNA polymerase may be subject to regulation through protein-protein interactions between the two ␣ subunits and two different species of transcription factors.The RNA polymerase holoenzyme of Escherichia coli is composed of core enzyme with subunit structure ␣ 2 ␤␤Ј, responsible for RNA polymerization, and one of multiple species of subunit, responsible for promoter recognition. Promoter selectivity of the holoenzyme is modulated by direct or indirect interaction with many transcription factors, resulting in switching of the global pattern of gene transcription according to the environment. The best-characterized target on the RNA polymerase involved in molecular communication with transcription factors is the ␣ subunit carboxy-terminal domain (CTD) that contains the contact sites for class I transcription factors. The ␣ subunit, consisting of 329 amino acid residues, is composed of two structural domains, each responsible for distinct functions (1-3) and each forming independent structural domains connected by a protease-sensitive flexible linker (4-6). The amino (N)-terminal domain from residues 20 to 235 plays a key role in RNA polymerase assembly by providing the contact surface for ␣ dimerization and binding of ␤ and ␤Ј subunits (7-10), whereas the CTD from residues 235 to 329 plays a regulatory role by providing the contact surfaces for trans-acting protein factors and cis-acting DNA elements (11)(12)(13)(14).Whereas the regulation of many E. coli promoters involves a single factor, some promoters are regulated by two or more transcription factors, and such coregulation systems involving multiple species of transcription factors can couple gene expression to diverse environmental conditions. Knowledge of the molecular mechanism of prokaryotic transcription regulation involving more than two factors would contribute much to understanding of the events carried out in eukaryotes, because the regulation of gene transcription in eukaryotes generally involves the action of multiple transcription factors. To gain insight into this problem, we analyzed interactions between RNA polymerase and cAMP receptor protein (CRP) dimers on promoters carrying tandem CRP-binding sites at various positions relative to the transcription start site. A set of promoters was constructed carrying one DNA site for CRP centered at position Ϫ41.5 upstream from the transcription start point and a second DNA site for CRP located further upstream (refs. 15

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Section: Resultssupporting

confidence: 85%

Positioning of two alpha subunit carboxy-terminal domains of RNA polymerase at promoters by two transcription factors

Murakami

Owens

Belyaeva

et al. 1997

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

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“…In E. coli, the CRPdependent promoters are grouped into three classes. Class I has a single CRP-binding site, centred at one of the positions 261?5, 271?5, 281?5 or 291?5; class II possesses a CRPrecognition site centred at 241?5, which thus overlaps with the 235 sequence; and class III requires two or more CRPbinding sites in the promoter for full activation (Busby & Ebright, 1999;Ebright, 1993;Ushida & Aiba, 1990). Based on the position of the CRP-recognition sequence 258?5 nt upstream of the TSP, the M. tuberculosis whiB1 promoter region most resembles the class I CRP-dependent promoters of E. coli.…”

Section: Discussionmentioning

confidence: 99%

Regulation of the expression of whiB1 in Mycobacterium tuberculosis: role of cAMP receptor protein

2006

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The wbl (whiB-like) genes encode putative transcription factors unique to actinomycetes. This study characterized the promoter element of one of the seven wbl genes of Mycobacterium tuberculosis, whiB1 (Rv3219c). The results reveal that whiB1 is transcribed by a class I-type cAMP receptor protein (CRP)-dependent promoter, harbouring a CRP-binding site positioned at "58?5 with respect to its transcription start point. In vivo promoter activity analysis and electrophoretic mobility shift assays suggest that the expression of whiB1 is indeed regulated by cAMP-dependent binding of CRP M (encoded by the M. tuberculosis gene Rv3676) to the whiB1 59 untranslated region (59UTR). b-Galactosidase gene fusion analysis revealed induction of the whiB1 promoter in M. tuberculosis on addition of exogenous dibutyric cAMP (a diffusible cAMP analogue) only when an intact CRP-binding site was present. These results indicate that M. tuberculosis whiB1 transcription is regulated in part by cAMP levels via direct binding of cAMP-activated CRP M to a consensus CRP-binding site in the whiB1 59UTR. INTRODUCTIONMycobacterium tuberculosis, the aetiological agent of tuberculosis, accounts for nearly 2 million human deaths every year. Approximately one-third of the world's population is latently infected with M. tuberculosis, and 7 to 8 million new TB cases occur annually (Dye et al., 1999). M. tuberculosis can survive in diverse surroundings ranging from the human host to droplet nuclei in the atmosphere. Adaptation to such diverse conditions requires controlled regulation of the expression of key genes that allow the bacillus to alter its physiology in response to changes in environmental stimuli. Sequencing of the M. tuberculosis genome has revealed the presence of more than 100 regulatory proteins, 13 sigma factors and 11 two-component systems (Cole et al., 1998), which provide the bacterium with a high degree of adaptability.The Wbl (WhiB-like) family of proteins is present throughout the actinomycetes but absent from all other organisms evaluated so far (Molle et al., 2000;Soliveri et al., 2000). Due to the presence of a conserved helix-turn-helix motif, these proteins are believed to function as DNA-binding transcription regulators. The first of these proteins, WhiB, was identified in Streptomyces coelicolor, a Gram-positive sporulating bacterium closely related to M. tuberculosis. S. coelicolor whiB mutants produce abnormally long, tightly coiled aerial hyphae that are completely blocked in their ability to form sporulation septa (Chater, 1972;Davis & Chater, 1992;Flardh et al., 1999). Studies of whiB orthologues in mycobacteria have shown that the M. smegmatis whiB2 gene (also called whmD) is essential ; M. tuberculosis whiB3 plays a role in virulence and its gene product may interact with a sigma factor of RNA polymerase (Steyn et al., 2002); whiB7 of M. tuberculosis is involved in multi-drug resistance (Morris et al., 2005). Each of the Wbl family of proteins contains four invariant cysteine residues, which are believed to be inv...

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“…1B). The CTDs of the two α subunits are attached to RNAP by flexible linkers, allowing these domains to make activatory contacts with proteins positioned up to a few tens of base pairs upstream of the promoter (25,26). The αCTDs can also bind directly to DNA sequences termed UP elements (27), which have been shown to stimulate transcription if located just upstream of the promoter (28).…”

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confidence: 99%

Enhancer-like long-range transcriptional activation by λ CI-mediated DNA looping

Cui

Murchland

Shearwin

et al. 2013

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

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How distant enhancer elements regulate the assembly of a transcription complex at a promoter remains poorly understood. Here, we use long-range gene regulation by the bacteriophage λ CI protein as a powerful system to examine this process in vivo. A 2.3-kb DNA loop, formed by CI bridging its binding sites at OR and OL, is known already to enhance repression at the lysogenic promoter PRM, located at OR. Here, we show that CI looping also activates PRM by allowing the C-terminal domain of the α subunit of the RNA polymerase bound at PRM to contact a DNA site adjacent to the distal CI sites at OL. Our results establish OL as a multifaceted enhancer element, able to activate transcription from long distances independently of orientation and position. We develop a physicochemical model of our in vivo data and use it to show that the observed activation is consistent with a simple recruitment mechanism, where the α-C-terminal domain to DNA contact need only provide ∼2.7 kcal/mol of additional binding energy for RNA polymerase. Structural modeling of this complete enhancer-promoter complex reveals how the contact is achieved and regulated, and suggests that distal enhancer elements, once appropriately positioned at the promoter, can function in essentially the same way as proximal promoter elements. I n eukaryotes, initiation of transcription is often regulated by enhancers, DNA sequences that can be located many kilobases away from the promoter. Enhancers are usually complex, being composed of multiple DNA elements that bind a variety of proteins (1, 2). The enhancer and the promoter are brought into close contact, looping the intervening DNA (3, 4), thus placing these binding elements close to the promoter. However, due to the lack of detailed understanding of enhancer-promoter complexes, how enhancers efficiently and specifically regulate initiation remains obscure (5).The action of DNA elements at and around a promoter is easier to understand. These sequences can be envisaged as a local scaffold for the assembly of transcription-favoring or transcription-inhibiting protein complexes. The promoter elements bind the transcription machinery, and promoter-proximal DNA elements position other proteins so that they can interact, favorably or unfavorably, with this machinery (2, 6-9). In prokaryotes, these promoter complexes are relatively simple. Repression of transcription is usually achieved by proteins positioned to compete with binding of RNA polymerase (RNAP), whereas transcription can be activated by proteins positioned adjacent to RNAP that contact it to stabilize initiation intermediates (8,10,11).The CI repressor of bacteriophage λ has been seminal in understanding how promoter-proximal elements are used in transcriptional regulation. Two CI binding sites, OR1 and OR2, overlap the lytic PR promoter and cooperative binding of two CI dimers to these sites competes with RNAP binding and represses the promoter. Extensive study of this complex has culminated in crystal structures that reveal how CI binds to the DNA a...

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Helical phase dependent action of CRP: effect of the distance between the CRP site and the −35 region on promoter activity

Cited by 141 publications

References 51 publications

Positioning of two alpha subunit carboxy-terminal domains of RNA polymerase at promoters by two transcription factors

Positioning of two alpha subunit carboxy-terminal domains of RNA polymerase at promoters by two transcription factors

Regulation of the expression of whiB1 in Mycobacterium tuberculosis: role of cAMP receptor protein

Enhancer-like long-range transcriptional activation by λ CI-mediated DNA looping

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