scholarly journals The bacterial multidrug resistance regulator BmrR distorts promoter DNA to activate transcription

2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Chengli Fang ◽  
Linyu Li ◽  
Yihan Zhao ◽  
Xiaoxian Wu ◽  
Steven J. Philips ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Multidrug Resistance ◽  
Rna Polymerase ◽  
Transcriptional Activation ◽  
Promoter Activity ◽  
Transcription Activation ◽  
Core Enzyme ◽  
Upstream Promoter ◽  
Promoter Dna

AbstractThe MerR-family proteins represent a unique family of bacteria transcription factors (TFs), which activate transcription in a manner distinct from canonical ones. Here, we report a cryo-EM structure of a B. subtilis transcription activation complex comprising B. subtilis six-subunit (2αββ‘ωε) RNA Polymerase (RNAP) core enzyme, σA, a promoter DNA, and the ligand-bound B. subtilis BmrR, a prototype of MerR-family TFs. The structure reveals that RNAP and BmrR recognize the upstream promoter DNA from opposite faces and induce four significant kinks from the −35 element to the −10 element of the promoter DNA in a cooperative manner, which restores otherwise inactive promoter activity by shortening the length of promoter non-optimal −35/−10 spacer. Our structure supports a DNA-distortion and RNAP-non-contact paradigm of transcriptional activation by MerR TFs.

scholarly journals Stepwise Recruitment of Components of the Preinitiation Complex by Upstream Activators In Vivo

1998 ◽  
Vol 18 (5) ◽  
pp. 2876-2883 ◽  
Author(s):  
Song He ◽  
Steven Jay Weintraub
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Mechanism Of Action ◽  
Transcriptional Activation ◽  
Tata Binding Protein ◽  
Preinitiation Complex ◽  
Functional Assays ◽  
Rna Polymerase Ii Holoenzyme

ABSTRACT Recently, it was found that if either the TATA binding protein or RNA polymerase II holoenzyme is artificially tethered to a promoter, transcription is activated. This finding provided presumptive evidence that upstream activating proteins function by recruiting components of the preinitiation complex (PIC) to the promoter. To date, however, there have been no studies demonstrating that upstream factors actually recruit components of the PIC to the promoter in vivo. Therefore, we have studied the mechanism of action of two disparate transactivating domains. We present a series of in vivo functional assays that demonstrate that each of these proteins targets different components of the PIC for recruitment. We show that, by targeting different components of the PIC for recruitment, these activating domains can cooperate with each other to activate transcription synergistically and that, even within one protein, two different activating subdomains can activate transcription synergistically by cooperating to recruit different components of the PIC. Finally, considering our work together with previous studies, we propose that certain transcription factors both recruit components of the PIC and facilitate steps in transcriptional activation that occur subsequent to recruitment.

Transcriptional takeover by σ appropriation: remodelling of the σ 70 subunit of Escherichia coli RNA polymerase by the bacteriophage T4 activator MotA and co-activator AsiA

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 1729-1740 ◽  
Author(s):  
Deborah M. Hinton ◽  
Suchira Pande ◽  
Neelowfar Wais ◽  
Xanthia B. Johnson ◽  
Madhavi Vuthoori ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Molecular Level ◽  
Bacteriophage T4 ◽  
Good Match ◽  
Promoter Sequences ◽  
Recognition Element ◽  
Upstream Promoter ◽  
Promoter Specificity ◽  
Promoter Dna

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 σ 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 σ 70 DNA element located in the −10 region. However, instead of the σ 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 σ 70 with the −10 region. This type of activation, which is called ‘σ appropriation’, is fundamentally different from other well-characterized models of prokaryotic activation in which an activator frequently serves to force σ 70 to contact a less than ideal −35 DNA element. This review summarizes the interactions of AsiA and MotA with σ 70, and discusses how these interactions accomplish the switch to T4 middle promoters by inhibiting the typical contacts of the C-terminal region of σ 70, region 4, with the host −35 DNA element and with other subunits of polymerase.

Investigations of the modular structure of bacterial promoters

2006 ◽  
Vol 73 ◽  
pp. 1-10 ◽  
Author(s):  
Nora S. Miroslavova ◽  
Stephen J.W. Busby
Keyword(s):  
Rna Polymerase ◽  
Dna Sequence ◽  
Transcription Initiation ◽  
Promoter Activity ◽  
Modular Structure ◽  
Bacterial Rna ◽  
Sequence Elements ◽  
Promoter Dna ◽  
Transcript Initiation ◽  
Rna Polymerase Holoenzyme

Bacterial RNA polymerase holoenzyme carries different determinants that contact different promoter DNA sequence elements. These contacts are essential for the recognition of promoters prior to transcript initiation. Here, we have investigated how active promoters can be built from different combinations of elements. Our results show that the contribution of different contacts to promoter activity is critically dependent on the overall promoter context, and that certain combinations of contacts can hinder transcription initiation.

scholarly journals Structure of the unique tetrameric STENOFOLIA homeodomain bound with target promoter DNA

2021 ◽  
Vol 77 (8) ◽  
Author(s):  
Prabhat Kumar Pathak ◽  
Fei Zhang ◽  
Shuxia Peng ◽  
Lifang Niu ◽  
Juhi Chaturvedi ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Transcriptional Activation ◽  
Hydrophobic Surface ◽  
Promoter Sequence ◽  
In Planta ◽  
Embryo Patterning ◽  
Stem Cell Maintenance ◽  
C Terminus ◽  
Promoter Dna

Homeobox transcription factors are key regulators of morphogenesis and development in both animals and plants. In plants, the WUSCHEL-related homeobox (WOX) family of transcription factors function as central organizers of several developmental programs ranging from embryo patterning to meristematic stem-cell maintenance through transcriptional activation and repression mechanisms. The Medicago truncatula STENOFOLIA (STF) gene is a master regulator of leaf-blade lateral development. Here, the crystal structure of the homeodomain (HD) of STF (STF-HD) in complex with its promoter DNA is reported at 2.1 Å resolution. STF-HD binds DNA as a tetramer, enclosing nearly the entire bound DNA surface. The STF-HD tetramer is partially stabilized by docking of the C-terminal tail of one protomer onto a conserved hydrophobic surface on the head of another protomer in a head-to-tail manner. STF-HD specifically binds TGA motifs, although the promoter sequence also contains TAAT motifs. Helix α3 not only serves a canonical role as a base reader in the major groove, but also provides DNA binding in the minor groove through basic residues located at its C-terminus. The structural and functional data in planta reported here provide new insights into the DNA-binding mechanisms of plant-specific HDs from the WOX family of transcription factors.

scholarly journals A highly conserved domain of RNA polymerase II shares a functional element with acidic activation domains of upstream transcription factors

1994 ◽  
Vol 14 (11) ◽  
pp. 7507-7516
Author(s):  
H Xiao ◽  
J D Friesen ◽  
J T Lis
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Normal Function ◽  
Yeast Saccharomyces Cerevisiae ◽  
Activation Domains ◽  
Acidic Domain ◽  
General Transcription Factors ◽  
Derivatives Of

We report here that the largest subunit of yeast RNA polymerase II contains an acidic domain that is similar to acidic activators of transcription. This domain includes the highly conserved homology box H. A hybrid protein containing this acidic domain fused to the DNA-binding domain of GAL4 is a potent activator of transcription in the yeast Saccharomyces cerevisiae. Interestingly, mutations that reduce the upstream activating activity of this acidic domain also abolish the normal function of RNA polymerase II. Such functional defects can be rescued by the acidic activation domains of VP16 and GAL4 when inserted into the mutant derivatives of RNA polymerase II. We further show that this acidic domain of RNA polymerase II interacts directly with two general transcription factors, the TATA-binding protein and TFIIB, and that the acidic activation domain of VP16 can compete specifically with the acidic domain of the RNA polymerase for these interactions. We discuss the implications of this finding for the mechanisms of transcriptional activation in eucaryotes.

scholarly journals A highly conserved domain of RNA polymerase II shares a functional element with acidic activation domains of upstream transcription factors.

1994 ◽  
Vol 14 (11) ◽  
pp. 7507-7516 ◽  
Author(s):  
H Xiao ◽  
J D Friesen ◽  
J T Lis
Keyword(s):  
Transcription Factors ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcriptional Activation ◽  
Normal Function ◽  
Yeast Saccharomyces Cerevisiae ◽  
Activation Domains ◽  
Acidic Domain ◽  
General Transcription Factors ◽  
Derivatives Of

We report here that the largest subunit of yeast RNA polymerase II contains an acidic domain that is similar to acidic activators of transcription. This domain includes the highly conserved homology box H. A hybrid protein containing this acidic domain fused to the DNA-binding domain of GAL4 is a potent activator of transcription in the yeast Saccharomyces cerevisiae. Interestingly, mutations that reduce the upstream activating activity of this acidic domain also abolish the normal function of RNA polymerase II. Such functional defects can be rescued by the acidic activation domains of VP16 and GAL4 when inserted into the mutant derivatives of RNA polymerase II. We further show that this acidic domain of RNA polymerase II interacts directly with two general transcription factors, the TATA-binding protein and TFIIB, and that the acidic activation domain of VP16 can compete specifically with the acidic domain of the RNA polymerase for these interactions. We discuss the implications of this finding for the mechanisms of transcriptional activation in eucaryotes.

scholarly journals Conformational properties of Bacillus subtilis RNA polymerase σA factor during transcription initiation

1993 ◽  
Vol 294 (1) ◽  
pp. 43-47 ◽  
Author(s):  
B Y Chang ◽  
R H Doi
Keyword(s):  
Rna Polymerase ◽  
Transcription Initiation ◽  
Trypsin Digestion ◽  
Tryptic Fragment ◽  
The Core ◽  
Core Enzyme ◽  
Conformational Properties ◽  
Promoter Dna ◽  
Rna Polymerase Holoenzyme ◽  
Dna Complex

By the use of a partial proteolysis method and Western-blot analysis, the conformational properties of Bacillus subtilis sigma A factor in the transcription initiation stage were studied. From a comparison of the trypsin-digestion patterns of free sigma A and of sigma A associated with core enzyme, it was found that the production of 45 kDa sigma A tryptic-derived fragment was enhanced when sigma A was associated with the core enzyme. More importantly, a 40 kDa sigma A tryptic-derived fragment was found exclusively in this associated state. Based on the change of the digestion kinetics when producing the 45 kDa tryptic fragment and the generation of this new 40 kDa tryptic fragment from sigma A, it was apparent that a conformation change of sigma A occurred during the association of sigma A with the core enzyme. Also, similar patterns were found for the sigma A present in the holoenzyme-promoter DNA complex. These findings suggest that no further distinctive conformational change of sigma A occurs at the step of RNA polymerase holoenzyme and promoter DNA complex formation. Trypsin-digestion patterns of sigma A in different RNA polymerase holoenzyme and promoter DNA complexes were also studied. The presence of similar trypsin digestion-patterns of sigma A in those complexes strongly supports the idea that a similar sigma A conformation is used in the recognition of different sigma A-type promoters and the formation of different open complexes.

scholarly journals Transcriptional activation of human 12-lipoxygenase gene promoter is mediated through Sp1 consensus sites in A431 cells

1997 ◽  
Vol 324 (1) ◽  
pp. 133-140 ◽  
Author(s):  
Yi-Wen LIU ◽  
Toshiya ARAKAWA ◽  
Shozo YAMAMOTO ◽  
Wen-Chang CHANG
Keyword(s):  
Transcription Factors ◽  
Transcriptional Activation ◽  
Binding Sites ◽  
Transcription Initiation ◽  
Promoter Activity ◽  
Start Codon ◽  
A431 Cells ◽  
Lipoxygenase Gene ◽  
Flanking Region ◽  
12 Lipoxygenase

The functional 5′ flanking region of the human 12-lipoxygenase in epidermoid carcinoma A431 cells was characterized. By a primer extension method, the transcription initiation sites were mapped at -47 adenosine, -48 guanosine and -55 guanosine upstream of the ATG translation start codon. Transient transfection with a series of 5′ and 3′ deletion constructs showed that the 5′ flanking region spanning from -224 to -100 bp was important for the basal expression of 12-lipoxygenase gene. Gel mobility shift assays with antibodies of transcription factors showed that both Sp1 and Sp3 required highly GC-rich Sp1 sites within this region for binding. Disruption of two Sp1 recognition motifs residing at -158 to -150 bp and -123 to -114 bp by site-directed mutagenesis markedly reduced the basal 12-lipoxygenase promoter activity and abolished the retarded bands in a gel-shift assay, indicating that these two Sp1-binding sites were essential for gene expression. The same two Sp1-binding sites in this promoter region were also responsible for epidermal growth factor (EGF)-induced expression of 12-lipoxygenase gene. Moreover, EGF also induced the transcriptional activation of luciferase driven by SV40 early promoter, which contained rich Sp1-binding sites. Taken together, the results suggest that two specific Sp1 consensus sites are involved in the mediation of the basal promoter activity as well as EGF induction of the 12-lipoxygenase gene and that Sp1 and Sp3 transcription factors might have a role in their regulation.

scholarly journals Spo0A Mutants of Bacillus subtilis with Sigma Factor-Specific Defects in Transcription Activation

1998 ◽  
Vol 180 (14) ◽  
pp. 3584-3591 ◽  
Author(s):  
Janet K. Hatt ◽  
Philip Youngman
Keyword(s):  
Bacillus Subtilis ◽  
Rna Polymerase ◽  
Transcriptional Activation ◽  
Sigma Factor ◽  
Transcription Activation ◽  
Sigma H ◽  
Rna Polymerase Holoenzyme ◽  
Amino Acid Segment ◽  
Identification And Characterization

ABSTRACT The transcription factor Spo0A of Bacillus subtilis has the unique ability to activate transcription from promoters that require different forms of RNA polymerase holoenzyme. One class of Spo0A-activated promoter, which includes spoIIEp, is recognized by RNA polymerase associated with the primary sigma factor, sigma A (ςA); the second, which includesspoIIAp, is recognized by RNA polymerase associated with an early-sporulation sigma factor, sigma H (ςH). Evidence suggests that Spo0A probably interacts directly with RNA polymerase to activate transcription from these promoters. To identify residues of Spo0A that may be involved in transcriptional activation, we used PCR mutagenesis of the entire spo0A gene and designed a screen using two distinguishable reporter fusions, spoIIE-gus andspoIIA-lacZ. Here we report the identification and characterization of five mutants of Spo0A that are specifically defective in activation of ςA-dependent promoters while maintaining activation of ςH-dependent promoters. These five mutants identify a 14-amino-acid segment of Spo0A, from residue 227 to residue 240, that is required for transcriptional activation of ςA-dependent promoters. This region may define a surface or domain of Spo0A that makes direct contacts with ςA-associated holoenzyme.

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