scholarly journals Crl activates transcription by stabilizing active conformation of the master stress transcription initiation factor

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Juncao Xu ◽  
Kaijie Cui ◽  
Liqiang Shen ◽  
Jing Shi ◽  
Lingting Li ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Transcription Initiation ◽  
Transcription Activation ◽  
Initiation Factor ◽  
Bacterial Cells ◽  
E Coli ◽  
Exchange Mass Spectrometry ◽  
Transcription Initiation Factor ◽  
Hydrogen Deuterium ◽  
Promoter Dna

σS is a master transcription initiation factor that protects bacterial cells from various harmful environmental stresses including antibiotic pressure. Although its mechanism remains unclear, it is known that full activation of σS-mediated transcription requires a σS-specific activator, Crl. In this study, we determined a 3.80 Å cryo-EM structure of an Escherichia coli transcription activation complex (E. coli Crl-TAC) comprising E. coli σS-RNA polymerase (σS-RNAP) holoenzyme, Crl, and a nucleic-acid scaffold. The structure reveals that Crl interacts with domain 2 of σS (σS2) and the RNAP core enzyme, but does not contact promoter DNA. Results from subsequent hydrogen-deuterium exchange mass spectrometry (HDX-MS) indicate that Crl stabilizes key structural motifs within σS2 to promote the assembly of the σS-RNAP holoenzyme and also to facilitate formation of an RNA polymerase–promoter DNA open complex (RPo). Our study demonstrates a unique DNA contact-independent mechanism of transcription activation, thereby defining a previously unrecognized mode of transcription activation in cells.

scholarly journals Crl activates transcription by stabilizing the active conformation of the master stress factor σS

2019 ◽  
Author(s):  
Juncao Xu ◽  
Kaijie Cui ◽  
Liqiang Shen ◽  
Jing Shi ◽  
Lingting Li ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Transcription Activation ◽  
Initiation Factor ◽  
Bacterial Cells ◽  
E Coli ◽  
Exchange Mass Spectrometry ◽  
Transcription Initiation Factor ◽  
Hydrogen Deuterium ◽  
Promoter Dna ◽  
Hdx Ms

SUMMARYσS is a master transcription initiation factor that protects bacterial cells from various harmful environmental stresses and antibiotic pressure. Although its mechanism remains unclear, it is known that full activation of σS-mediated transcription requires a σS-specific activator, Crl. In this study, we determined a 3.80 Å cryo-EM structure of an E. coli transcription activation complex (E. coli Crl-TAC) comprising E. coli σS-RNAP holoenzyme, Crl, and a nucleic-acid scaffold. The structure reveals that Crl interacts with the domain 2 of σS (σS2), sharing no interaction with promoter DNA. Subsequent hydrogen-deuterium exchange mass spectrometry (HDX-MS) results indicate that Crl stabilizes key structural motifs of σS2 to promote the assembly of σS-RNAP holoenzyme and also to facilitate formation of the RNA polymerase-promoter DNA open complex (RPo). Our study demonstrates a unique DNA contact-independent mechanism of transcription activation, thereby defining a previously unrecognized mode of transcription activation in cells.

scholarly journals Inhibition of TATA Binding Protein Dimerization by RNA Polymerase III Transcription Initiation Factor Brf1

2004 ◽  
Vol 279 (31) ◽  
pp. 32401-32406 ◽  
Author(s):  
Diane E. Alexander ◽  
David J. Kaczorowski ◽  
Amy J. Jackson-Fisher ◽  
Drew M. Lowery ◽  
Sara J. Zanton ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Transcription Initiation ◽  
Binding Protein ◽  
Rna Polymerase Iii ◽  
Tata Binding Protein ◽  
Initiation Factor ◽  
Protein Dimerization ◽  
Polymerase Iii ◽  
Transcription Initiation Factor

scholarly journals Np4A alarmones function in bacteria as precursors to RNA caps

2020 ◽  
Vol 117 (7) ◽  
pp. 3560-3567 ◽  
Author(s):  
Daniel J. Luciano ◽  
Joel G. Belasco
Keyword(s):  
Rna Polymerase ◽  
Transcription Initiation ◽  
Promoter Sequence ◽  
Rna Degradation ◽  
Initiation Site ◽  
Bacterial Cells ◽  
E Coli ◽  
N Incorporation ◽  
Nascent Transcripts

Stresses that increase the cellular concentration of dinucleoside tetraphosphates (Np4Ns) have recently been shown to impact RNA degradation by inducing nucleoside tetraphosphate (Np4) capping of bacterial transcripts. However, neither the mechanism by which such caps are acquired nor the function of Np4Ns in bacteria is known. Here we report that promoter sequence changes upstream of the site of transcription initiation similarly affect both the efficiency with which Escherichia coli RNA polymerase incorporates dinucleoside polyphosphates at the 5′ end of nascent transcripts in vitro and the percentage of transcripts that are Np4-capped in E. coli, clear evidence for Np4 cap acquisition by Np4N incorporation during transcription initiation in bacterial cells. E. coli RNA polymerase initiates transcription more efficiently with Np4As than with ATP, particularly when the coding strand nucleotide that immediately precedes the initiation site is a purine. Together, these findings indicate that Np4Ns function in bacteria as precursors to Np4 caps and that RNA polymerase has evolved a predilection for synthesizing capped RNA whenever such precursors are abundant.

scholarly journals E. coli TraR allosterically regulates transcription initiation by altering RNA polymerase conformation and dynamics

2019 ◽  
Author(s):  
James Chen ◽  
Saumya Gopalkrishnan ◽  
Courtney Chiu ◽  
Albert Y. Chen ◽  
Elizabeth A. Campbell ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Conformational Changes ◽  
Transcription Initiation ◽  
Structural Changes ◽  
E Coli ◽  
Bacterial Proteins ◽  
Cryo Electron Microscopy ◽  
Promoter Complex ◽  
Promoter Dna ◽  
Induced Changes

AbstractTraR and its homolog DksA are bacterial proteins that regulate transcription initiation by binding directly to RNA polymerase (RNAP) rather than to promoter DNA. Effects of TraR mimic the combined effects of DksA and its cofactor ppGpp. How TraR and its homologs regulate transcription is unclear. Here, we use cryo-electron microscopy to determine structures of Escherichia coli RNAP, with or without TraR, and of an RNAP-promoter complex. TraR binding induced RNAP conformational changes not seen in previous crystallographic analyses, and a quantitative analysis of RNAP conformational heterogeneity revealed TraR-induced changes in RNAP dynamics. These changes involve mobile regions of RNAP affecting promoter DNA interactions, including the βlobe, the clamp, the bridge helix, and several lineage-specific insertions. Using mutational approaches, we show that these structural changes, as well as effects on σ70 region 1.1, are critical for transcription activation or inhibition, depending on the kinetic features of regulated promoters.

The C-terminal domain of the largest subunit of RNA polymerase II and transcription initiation

1989 ◽  
Vol 9 (12) ◽  
pp. 5750-5753
Author(s):  
M Moyle ◽  
J S Lee ◽  
W F Anderson ◽  
C J Ingles
Keyword(s):  
Monoclonal Antibodies ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Transcription Initiation ◽  
Initiation Factor ◽  
Transcription Initiation Factor ◽  
Evolutionarily Conserved ◽  
Repeat Domain ◽  
Initiation Of Transcription

Monoclonal antibodies specific for the evolutionarily conserved C-terminal heptapeptide repeat domain of the largest subunit of RNA polymerase II inhibited the initiation of transcription from mammalian promoters in vitro. Since these antibodies did not inhibit elongation and randomly initiated transcription, the heptapeptide repeats may function by binding class II transcription initiation factor(s).

scholarly journals Function of the growth-regulated transcription initiation factor TIF-IA in initiation complex formation at the murine ribosomal gene promoter.

1993 ◽  
Vol 13 (11) ◽  
pp. 6723-6732 ◽  
Author(s):  
A Schnapp ◽  
G Schnapp ◽  
B Erny ◽  
I Grummt
Keyword(s):  
Cell Proliferation ◽  
Rna Polymerase ◽  
Transcription Initiation ◽  
Human Cells ◽  
Initiation Factor ◽  
Rrna Genes ◽  
Regulation Of Transcription ◽  
Initiation Complex ◽  
Transcription System ◽  
Transcription Initiation Factor

Alterations in the rate of cell proliferation are accompanied by changes in the transcription of rRNA genes. In mammals, this growth-dependent regulation of transcription of genes coding for rRNA (rDNA) is due to reduction of the amount or activity of an essential transcription factor, called TIF-IA. Extracts prepared from quiescent cells lack this factor activity and, therefore, are transcriptionally inactive. We have purified TIF-IA from exponentially growing cells and have shown that it is a polypeptide with a molecular mass of 75 kDa which exists as a monomer in solution. Using a reconstituted transcription system consisting of purified transcription factors, we demonstrate that TIF-IA is a bona fide transcription initiation factor which interacts with RNA polymerase I. Preinitiation complexes can be assembled in the absence of TIF-IA, but formation of the first phosphodiester bonds of nascent rRNA is precluded. After initiation, TIF-IA is liberated from the initiation complex and facilitates transcription from templates bearing preinitiation complexes which lack TIF-IA. Despite the pronounced species specificity of class I gene transcription, this growth-dependent factor has been identified not only in mouse but also in human cells. Murine TIF-IA complements extracts from both growth-inhibited mouse and human cells. The analogous human activity appears to be similar or identical to that of TIF-IA. Therefore, despite the fact that the RNA polymerase transcription system has evolved sufficiently rapidly that an rDNA promoter from one species will not function in another species, the basic mechanisms that adapt ribosome synthesis to cell proliferation have been conserved.

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