scholarly journals Regulation of Gene Expression of phiEco32-like Bacteriophage 7-11

2022 ◽  
Author(s):  
Daria Lavysh ◽  
Vladimir Mekler ◽  
Evgeny Klimuk ◽  
Konstantin Severinov
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Regulation Of Gene Expression ◽  
In Vitro Transcription ◽  
Single Element ◽  
Bacterial Rna ◽  
Sigma 70 ◽  
Rna Polymerase Holoenzyme ◽  
Rna Polymerase Promoter

Salmonella enterica serovar Newport bacteriophage 7-11 shares 41 homologous ORFs with Escherichia coli phage phiEco32 and both phages encode a protein similar to bacterial RNA polymerase promoter specificity  subunit. Here, we investigated the temporal pattern of 7-11 gene expression during the infection and compared it to the previously determined transcription strategy of phiEco32. Using primer extension and in vitro transcription assays we identified eight promoters recognized by host RNA polymerase holoenzyme containing 7-11  subunit SaPh711_gp47. These promoters are characterized by a bipartite consensus GTAAtg-(16)-aCTA and are located upstream of late phage genes. While dissimilar from single-element middle and late promoters of phiEco32 recognized by holoenzyme formed by the phi32_gp36  factor, the 7-11 late promoters are located at genome positions similar to those of phiEco32 middle and late promoters. Two early 7-11 promoters are recognized by RNA polymerase holoenzyme containing host primary σ70 factor. Unlike the case of phiEco32, no shut off of σ70-dependent transcription is observed during 7-11 infection and there are no middle promoters. These differences can be explained by the fact that phage 7-11 does not encode a homologue of phi32_gp79, an inhibitor of host and early phage transcription and an activator of transcription by the phi32_gp36-holoenzyme.

scholarly journals Recognition of Streptococcal Promoters by the Pneumococcal SigA Protein

2021 ◽  
Vol 8 ◽  
Author(s):  
Virtu Solano-Collado ◽  
Sofía Ruiz-Cruz ◽  
Fabián Lorenzo-Díaz ◽  
Radoslaw Pluta ◽  
Manuel Espinosa ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Sigma Factor ◽  
Regulation Of Gene Expression ◽  
E Coli ◽  
Core Enzyme ◽  
Protein Architecture ◽  
Bacterial Rna ◽  
Promoter Recognition ◽  
Rna Polymerase Holoenzyme

Promoter recognition by RNA polymerase is a key step in the regulation of gene expression. The bacterial RNA polymerase core enzyme is a complex of five subunits that interacts transitory with one of a set of sigma factors forming the RNA polymerase holoenzyme. The sigma factor confers promoter specificity to the RNA polymerase. In the Gram-positive pathogenic bacterium Streptococcus pneumoniae, most promoters are likely recognized by SigA, a poorly studied housekeeping sigma factor. Here we present a sequence conservation analysis and show that SigA has similar protein architecture to Escherichia coli and Bacillus subtilis homologs, namely the poorly conserved N-terminal 100 residues and well-conserved rest of the protein (domains 2, 3, and 4). Further, we have purified the native (untagged) SigA protein encoded by the pneumococcal R6 strain and reconstituted an RNA polymerase holoenzyme composed of the E. coli core enzyme and the sigma factor SigA (RNAP-SigA). By in vitro transcription, we have found that RNAP-SigA was able to recognize particular promoters, not only from the pneumococcal chromosome but also from the S. agalactiae promiscuous antibiotic-resistance plasmid pMV158. Specifically, SigA was able to direct the RNA polymerase to transcribe genes involved in replication and conjugative mobilization of plasmid pMV158. Our results point to the versatility of SigA in promoter recognition and its contribution to the promiscuity of plasmid pMV158.

scholarly journals Structural Organization of Bacterial RNA Polymerase Holoenzyme and the RNA Polymerase-Promoter Open Complex

Cell ◽  
2002 ◽  
Vol 108 (5) ◽  
pp. 599-614 ◽  
Author(s):  
Vladimir Mekler ◽  
Ekaterine Kortkhonjia ◽  
Jayanta Mukhopadhyay ◽  
Jennifer Knight ◽  
Andrei Revyakin ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Structural Organization ◽  
Bacterial Rna ◽  
Rna Polymerase Holoenzyme ◽  
Rna Polymerase Promoter

scholarly journals The Ferredoxin-Like Protein FerR Regulates PrbP Activity inLiberibacter asiaticus

2018 ◽  
Vol 85 (4) ◽  
Author(s):  
Lei Pan ◽  
Danilo da Silva ◽  
Fernando A. Pagliai ◽  
Natalie A. Harrison ◽  
Claudio F. Gonzalez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Rna Polymerase ◽  
Regulation Of Gene Expression ◽  
Accessory Protein ◽  
Mrna Levels ◽  
Plant Host ◽  
Content Type ◽  
Environmental Signals ◽  
Liberibacter Asiaticus

ABSTRACTInLiberibacter asiaticus, PrbP is an important transcriptional accessory protein that regulates gene expression through interactions with the RNA polymerase β-subunit and a specific sequence on the promoter region. The constitutive expression ofprbPobserved upon chemical inactivation of PrbP-DNA interactionsin vivoindicated that the expression ofprbPwas not autoregulated at the level of transcription. This observation suggested that a modulatory mechanism via protein-protein interactions may be involved.In silicogenome association analysis identified FerR (CLIBASIA_01505), a putative ferredoxin-like protein, as a PrbP-interacting protein. Using a bacterial two-hybrid system and immunoprecipitation assays, interactions between PrbP and FerR were confirmed.In vitrotranscription assays were used to show that FerR can increase the activity of PrbP by 16-fold when present in the PrbP-RNA polymerase reaction mixture. The FerR protein-protein interaction surface was predicted by structural modeling and followed by site-directed mutagenesis. Amino acids V20, V23, and C40 were identified as the most important residues in FerR involved in the modulation of PrbP activityin vitro. The regulatory mechanism of FerR abundance was examined at the transcription level. In contrast toprbPofL. asiaticus(prbPLas), mRNA levels offerRofL. asiaticus(ferRLas) are induced by an increase in osmotic pressure. The results of this study revealed that the activity of the transcriptional activator PrbPLasis modulated via interactions with FerRLas. The induction offerRLasexpression by osmolarity provides insight into the mechanisms of adjusting gene expression in response to host environmental signals inL. asiaticus.IMPORTANCEThe rapid spread and aggressive progression of huanglongbing (HLB) in the major citrus-producing areas have raised global recognition of and vigilance to this disease. As a result, the causative agent,Liberibacter asiaticus, has been investigated from various perspectives. However, gene expression regulatory mechanisms that are important for the survival and persistence of this intracellular pathogen remain largely unexplored. PrbP is a transcriptional accessory protein important forL. asiaticussurvival in the plant host. In this study, we investigated the interactions between PrbP inL. asiaticus(PrbPLas) and a ferredoxin-like protein (FerR) inL. asiaticus, FerRLas. We show that the presence of FerR stabilizes and augments the activity of PrbPLas. In addition, we demonstrate that the expression offerRis induced by increases in osmolarity inLiberibacter crescens. Altogether, these results suggest that FerRLasand PrbPLasmay play important roles in the regulation of gene expression in response to changing environmental signals duringL. asiaticusinfection in the citrus host.

scholarly journals Properties of Escherichia coli RNA polymerase from a strain devoid of the stringent response alarmone ppGpp.

2008 ◽  
Vol 55 (2) ◽  
pp. 317-323 ◽  
Author(s):  
Agnieszka Szalewska-Pałasz
Keyword(s):  
Escherichia Coli ◽  
Rna Polymerase ◽  
Stringent Response ◽  
In Vitro Transcription ◽  
Complex Stability ◽  
Wild Type ◽  
Guanosine Tetraphosphate ◽  
Sigma 70 ◽  
In Vitro Performance

The stringent response alarmone guanosine tetraphosphate (ppGpp) affects transcription from many promoters. ppGpp binds directly to the transcription enzyme of Escherichia coli, RNA polymerase. Analysis of the crystal structure of RNA polymerase with ppGpp suggested that binding of this nucleotide may result in some conformational or post-translational alterations to the enzyme. These changes might affect in vitro performance of the enzyme. Here, a comparison of the in vitro properties of RNA polymerases isolated from wild type and ppGpp-deficient bacteria shows that both enzymes do not differ in i) transcription activity of various promoters (e.g. sigma(70)-rrnB P1, lambdapL, T7A1), ii) response to ppGpp, iii) promoter-RNA polymerase open complex stability. Thus, it may be concluded that ppGpp present in the bacterial cell prior to purification of the RNA polymerase does not result in the alterations to the enzyme that could be permanent and affect its in vitro transcription capacity.

scholarly journals Bacteriophage SP6-specific RNA polymerase. II. Mapping of SP6 DNA and selective in vitro transcription.

1982 ◽  
Vol 257 (10) ◽  
pp. 5779-5788 ◽  
Author(s):  
G A Kassavetis ◽  
E T Butler ◽  
D Roulland ◽  
M J Chamberlin
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
In Vitro Transcription
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