Bipartite functional map of the E. coli RNA polymerase α subunit: Involvement of the C-terminal region in transcription activation by cAMP-CRP

Igarashi, Kazuhiko; Ishihama, Akira

doi:10.1016/0092-8674(91)90553-b

Cited by 342 publications

(350 citation statements)

References 32 publications

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“…Recombinant exuR was overexpressed from pGEME_his vector that was constructed on the basis of pGEMDXba (Igarashi & Ishihama, 1991) using primers exuR_beg and exuR_end_his (Table S2), with six histidine codons added on its 3¢-end. As the DNA-binding domain of ExuR is located on the N-terminal end, addition of His-tag did not affect its functionality.…”

Section: Methodsmentioning

confidence: 99%

Control of hexuronate metabolism in Escherichia coli by the two interdependent regulators, ExuR and UxuR: derepression by heterodimer formation

et al. 2016

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Section: Methodsmentioning

confidence: 99%

Control of hexuronate metabolism in Escherichia coli by the two interdependent regulators, ExuR and UxuR: derepression by heterodimer formation

et al. 2016

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“…The activities of the reconstituted RNA polymerases were assayed by in vitro transcription with linear tac and htrA DNAs as templates. As shown previously (8), in addition to the core RNA polymerase (E), 70 was required for specific transcription initiated from the tac promoter (Fig. 4, lanes 1 and 3).…”

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confidence: 95%

The rpoE gene of Escherichia coli, which encodes sigma E, is essential for bacterial growth at high temperature

et al. 1995

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In vitro transcription analysis has shown that only RNA polymerase containing an alternative sigma subunit, E , activates transcription from one of the rpoH promoters and the htrA promoter. The location of the rpoE gene encoding E on the Escherichia coli chromosome has recently been established, but no rpoE mutant has yet become available for phenotypic testing. We cloned the rpoE gene from the -ordered clones of the E. coli genome and confirmed that the reconstituted RNA polymerase containing the gene product (E E ) can transcribe htrA in vitro. We constructed an rpoE-defective strain by gene disruption using the cloned rpoE gene. We demonstrate that expression of htrA is completely dependent on the rpoE gene in vivo and that the rpoE gene is essential for bacterial growth at high temperature.In Escherichia coli, RNA polymerase consists of the subunits of 2␣, ␤, ␤Ј, and a specific . The subunit directs RNA polymerase to initiate transcription at promoter sites on the DNA (6). The primary factor, 70 encoded by rpoD, is responsible for transcription of the majority of the genes expressed during exponential growth. In addition, alternative factors direct transcription of sets of genes whose products are needed for specific functions, such as nitrogen fixation, flagellum synthesis, heat shock response, and stationary-phase growth (4).The activities of two alternative factors, 32 and E , increase after the cell is exposed to a high temperature or ethanol (2,3,5,21,25,28). RNA polymerase containing 32 (E 32 ) transcribes the heat shock genes whose products are related to chaperones and proteases (29). The functions of many of these heat shock proteins are involved in binding to cytoplasmic proteins and assist the cytoplasmic proteins in folding or unfolding (7,15,22,26) and in protecting the cell from severe stresses such as exposure to 10% ethanol or a 50ЊC temperature (18). Using an in vitro transcription assay, Erickson and Gross (2) and Wang and Kaguni (25) have independently identified the E subunit that is essential for transcription from one (rpoHp 3 ) of the promoters of rpoH encoding 32 and the promoter of htrA encoding a periplasmic endopeptidase essential for growth at high temperatures. Mecsas et al. (16) have proposed that the E regulon is involved in the processes that function in the extracytoplasmic compartments.

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“…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.…”

mentioning

confidence: 99%

“…The ␣ subunit, consisting of 329 amino acid residues, is composed of two structural domains, each responsible for distinct functions (1)(2)(3) and each forming independent structural domains connected by a protease-sensitive flexible linker (4)(5)(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)(8)(9)(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).…”

mentioning

confidence: 99%

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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Bipartite functional map of the E. coli RNA polymerase α subunit: Involvement of the C-terminal region in transcription activation by cAMP-CRP

Cited by 342 publications

References 32 publications

Control of hexuronate metabolism in Escherichia coli by the two interdependent regulators, ExuR and UxuR: derepression by heterodimer formation

Control of hexuronate metabolism in Escherichia coli by the two interdependent regulators, ExuR and UxuR: derepression by heterodimer formation

The rpoE gene of Escherichia coli, which encodes sigma E, is essential for bacterial growth at high temperature

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

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