Transcription of the Streptococcus pyogenes Hyaluronic Acid Capsule Biosynthesis Operon Is Regulated by Previously Unknown Upstream Elements

ABSTRACTThe important human pathogenStreptococcus pyogenes(group AStreptococcus[GAS]) produces a hyaluronic acid (HA) capsule that plays critical roles in immune evasion. Previous studies showed that thehasABCoperon encoding the capsule biosynthesis enzymes is under the control of a single promoter, P1, which is negatively regulated by the two-component regulatory system CovR/S. In this work, we characterize the sequence upstream of P1 and identify a novel regulatory region controlling transcription of the capsule biosynthesis operon in the M1 serotype strain MGAS2221. This region consists of a promoter, P2, which initiates transcription of a novel small RNA, HasS, an intrinsic transcriptional terminator that inefficiently terminates HasS, permitting read-through transcription ofhasABC, and a putative promoter which lies upstream of P2. Electrophoretic mobility shift assays, quantitative reverse transcription-PCR, and transcriptional reporter data identified CovR as a negative regulator of P2. We found that the P1 and P2 promoters are completely repressed by CovR, and capsule expression is regulated by the putative promoter upstream of P2. Deletion ofhasSor of the terminator eliminates CovR-binding sequences, relieving repression and increasing read-through,hasAtranscription, and capsule production. Sequence analysis of 44 GAS genomes revealed a high level of polymorphism in the HasS sequence region. Most of the HasS variations were located in the terminator sequences, suggesting that this region is under strong selective pressure. We discovered that the terminator deletion mutant is highly resistant to neutrophil-mediated killing and is significantly more virulent in a mouse model of GAS invasive disease than the wild-type strain. Together, these results are consistent with the naturally occurring mutations in this region modulating GAS virulence.

Download Full-text

CovR and VicRKX Regulate Transcription of the Collagen Binding Protein Cnm ofStreptococcus mutans

Journal of Bacteriology ◽

10.1128/jb.00141-18 ◽

2018 ◽

Vol 200 (23) ◽

Cited By ~ 2

Author(s):

Lívia Araújo Alves ◽

Tridib Ganguly ◽

Renata O. Mattos-Graner ◽

Jessica Kajfasz ◽

Erika N. Harth-Chu ◽

...

Keyword(s):

Streptococcus Mutans ◽

Bacterial Infections ◽

Regulatory Region ◽

Core Gene ◽

Negative Regulator ◽

Mobility Shift ◽

Content Type ◽

Intracellular Invasion ◽

Systemic Infections

ABSTRACTCnm is a surface-associated protein present in a subset ofStreptococcus mutansstrains that mediates binding to extracellular matrices, intracellular invasion, and virulence. Here, we showed thatcnmtranscription is controlled by the global regulators CovR and VicRKX.In silicoanalysis identified multiple putative CovR- and VicR-binding motifs in the regulatory region ofcnmas well as in the downstream genepgfS, which is associated with the posttranslational modification of Cnm. Electrophoretic mobility shift assays revealed that CovR and VicR specifically and independently bind to thecnmandpgfSpromoter regions. Quantitative real-time PCR and Western blot analyses of ΔcovRand ΔvicKstrains as well as of a strain overexpressingvicRKXrevealed that CovR functions as a positive regulator ofcnm, whereas VicRKX acts as a negative regulator. In agreement with the role of VicRKX as a repressor, the ΔvicKstrain showed enhanced binding to collagen and laminin and higher intracellular invasion rates. Overexpression ofvicRKXwas associated with decreased rates of intracellular invasion but did not affect collagen or lamin binding activities, suggesting that this system controls additional genes involved in binding to these extracellular matrix proteins. As expected, based on the role of CovR incnmregulation, the ΔcovRstrain showed decreased intracellular invasion rates, but, unexpectedly collagen and laminin binding activities were increased in this mutant strain. Collectively, the results presented here expand the repertoire of virulence-related genes regulated by CovR and VicRKX to include the core genepgfSand the noncore genecnm.IMPORTANCEStreptococcus mutansis a major pathogen associated with dental caries and also implicated in systemic infections, in particular, infective endocarditis. The Cnm adhesin ofS. mutansis an important virulence factor associated with systemic infections and caries severity. Despite its role in virulence, the regulatory mechanisms governingcnmexpression are poorly understood. Here, we describe the identification of two independent regulatory systems controlling the transcription ofcnmand the downstreampgfS-pgfM1-pgfE-pgfM2operon. A better understanding of the mechanisms controlling expression of virulence factors like Cnm can facilitate the development of new strategies to treat bacterial infections.

Download Full-text

Coordinate Control of the Locus of Enterocyte Effacement and Enterohemolysin Genes by Multiple Common Virulence Regulators in Enterohemorrhagic Escherichia coli

Infection and Immunity ◽

10.1128/iai.05023-11 ◽

2011 ◽

Vol 79 (11) ◽

pp. 4628-4637 ◽

Cited By ~ 11

Author(s):

Sunao Iyoda ◽

Naoko Honda ◽

Takehito Saitoh ◽

Ken Shimuta ◽

Jun Terajima ◽

...

Keyword(s):

Escherichia Coli ◽

Regulatory Region ◽

Regulatory System ◽

Enterohemorrhagic Escherichia Coli ◽

Mobility Shift ◽

Content Type ◽

Positive Regulators ◽

Enterocyte Effacement ◽

Gel Mobility Shift ◽

K 12

ABSTRACTThe locus of enterocyte effacement (LEE) pathogenicity island is required for the intimate adhesion of enterohemorrhagicEscherichia coli(EHEC) to the intestinal epithelial cells. GrlR and GrlA are LEE-encoded negative and positive regulators, respectively. The interaction of these two regulators is important for controlling the transcription of LEE genes through Ler, a LEE-encoded central activator for the LEE. The GrlR-GrlA regulatory system controls not only LEE but also the expression of the flagellar and enterohemolysin (Ehx) genes in EHEC. Since Ehx levels were markedly induced in agrlRmutant but not in agrlR grlAdouble mutant and significantly increased by overexpression of GrlA in alermutant, GrlA is responsible for this regulation (T. Saitoh et al., J. Bacteriol.190:4822-4830, 2008). In this study, additional investigations of the regulation ofehxgene expression determined that Ler also acts as an activator for Ehx expression without requiring GrlA function. We recently reported that the LysR-type regulator LrhA positively controls LEE expression (N. Honda et al., Mol. Microbiol.74:1393-1411, 2009). The hemolytic activity of thelrhAmutant strain of EHEC was lower than that of the wild-type strain, and LrhA markedly inducedehxtranscription in anE. coliK-12 strain, suggesting that LrhA also activates the transcription ofehxwithout GrlA and Ler. Gel mobility shift assays demonstrated that Ler and LrhA directly bind to the regulatory region ofehxC. Together, these results indicate that transcription ofehxis positively regulated by Ler, GrlA, and LrhA, which all act as positive regulators for LEE expression.

Download Full-text

The Novel Streptococcal Transcriptional Regulator XtgS Negatively Regulates Bacterial Virulence and Directly Represses PseP Transcription

Infection and Immunity ◽

10.1128/iai.00035-20 ◽

2020 ◽

Vol 88 (10) ◽

Author(s):

Guangjin Liu ◽

Tingting Gao ◽

Xiaojun Zhong ◽

Jiale Ma ◽

Yumin Zhang ◽

...

Keyword(s):

Strong Association ◽

Challenge Test ◽

Transcriptional Regulator ◽

Transcriptional Regulators ◽

Negative Regulator ◽

Broad Host Range ◽

Bacterial Survival ◽

Mobility Shift ◽

Upstream Gene ◽

Content Type

ABSTRACT Streptococcus agalactiae (group B streptococcus [GBS]) has received continuous attention for its involvement in invasive infections and its broad host range. Transcriptional regulators have an important impact on bacterial adaptation to various environments. Research on transcriptional regulators will shed new light on GBS pathogenesis. In this study, we identified a novel XRE-family transcriptional regulator encoded on the GBS genome, designated XtgS. Our data demonstrate that XtgS inactivation significantly increases bacterial survival in host blood and animal challenge test, suggesting that it is a negative regulator of GBS pathogenicity. Further transcriptomic analysis and quantitative reverse transcription-PCR (qRT-PCR) mainly indicated that XtgS significantly repressed transcription of its upstream gene pseP. Based on electrophoretic mobility shift and lacZ fusion assays, we found that an XtgS homodimer directly binds a palindromic sequence in the pseP promoter region. Meanwhile, the PseP and XtgS combination naturally coexists in diverse Streptococcus genomes and has a strong association with sequence type, serotype diversification and host adaptation of GBS. Therefore, this study reveals that XtgS functions as a transcriptional regulator that negatively affects GBS virulence and directly represses PseP expression, and it provides new insights into the relationships between transcriptional regulator and genome evolution.

Download Full-text

Polymyxin B Resistance and Biofilm Formation in Vibrio cholerae Are Controlled by the Response Regulator CarR

Infection and Immunity ◽

10.1128/iai.02700-14 ◽

2015 ◽

Vol 83 (3) ◽

pp. 1199-1209 ◽

Cited By ~ 38

Author(s):

Kivanc Bilecen ◽

Jiunn C. N. Fong ◽

Andrew Cheng ◽

Christopher J. Jones ◽

David Zamorano-Sánchez ◽

...

Keyword(s):

Vibrio Cholerae ◽

Biofilm Formation ◽

Lipid A ◽

Response Regulator ◽

Regulatory Region ◽

Regulatory System ◽

Polymyxin B ◽

Content Type ◽

Two Component ◽

Antimicrobial Peptide Resistance

Two-component systems play important roles in the physiology of many bacterial pathogens.Vibrio cholerae's CarRS two-component regulatory system negatively regulates expression ofvps(Vibriopolysaccharide) genes and biofilm formation. In this study, we report that CarR confers polymyxin B resistance by positively regulating expression of thealmEFGgenes, whose products are required for glycine and diglycine modification of lipid A. We determined that CarR directly binds to the regulatory region of thealmEFGoperon. Similarly to acarRmutant, strains lackingalmE,almF, andalmGexhibited enhanced polymyxin B sensitivity. We also observed that strains lackingalmEor thealmEFGoperon have enhanced biofilm formation. Our results reveal that CarR regulates biofilm formation and antimicrobial peptide resistance inV. cholerae.

Download Full-text

Complete Genome Sequence of Hypervirulent Streptococcus pyogenes emm3 Strain 1838

Microbiology Resource Announcements ◽

10.1128/mra.01494-18 ◽

2019 ◽

Vol 8 (2) ◽

Author(s):

Benfang Lei ◽

Anthony R. Flores ◽

Carl Yeoman ◽

Mengyao Liu

Keyword(s):

Genome Sequence ◽

Streptococcus Pyogenes ◽

Complete Genome Sequence ◽

Complete Genome ◽

Toxic Shock Syndrome ◽

Regulatory System ◽

Toxic Shock ◽

Content Type

We report the complete genome sequence for Streptococcus pyogenes strain 1838 (type emm3) isolated from a patient with toxic shock syndrome. The strain lacked the speK- and sla-encoding prophage frequently encountered among emm3 strains and possessed an Arg66His mutation in CovR of the 2-component virulence regulatory system CovRS.

Download Full-text

The Histone-Like Nucleoid Structuring Protein (H-NS) Is a Negative Regulator of the Lateral Flagellar System in the Deep-Sea Bacterium Shewanella piezotolerans WP3

Applied and Environmental Microbiology ◽

10.1128/aem.00297-16 ◽

2016 ◽

Vol 82 (8) ◽

pp. 2388-2398 ◽

Cited By ~ 6

Author(s):

Huahua Jian ◽

Guanpeng Xu ◽

Yingbao Gai ◽

Jun Xu ◽

Xiang Xiao

Keyword(s):

Deep Sea ◽

Negative Regulator ◽

Mobility Shift ◽

Swarming Motility ◽

Content Type ◽

Cold Environments ◽

Expression Of Genes ◽

Gel Mobility Shift ◽

Pressure Stress ◽

Flagellar Genes

ABSTRACTAlthough the histone-like nucleoid structuring protein (H-NS) is well known for its involvement in the adaptation of mesophilic bacteria, such asEscherichia coli, to cold environments and high-pressure stress, an understanding of the role of H-NS in the cold-adapted benthic microorganisms that live in the deep-sea ecosystem, which covers approximately 60% of the earth's surface, is still lacking. In this study, we characterized the function of H-NS inShewanella piezotoleransWP3, which was isolated from West Pacific sediment at a depth of 1,914 m. Anhnsgene deletion mutant (WP3Δhns) was constructed, and comparative whole-genome microarray analysis was performed. H-NS had a significant influence (fold change, >2) on the expression of a variety of WP3 genes (274 and 280 genes were upregulated and downregulated, respectively), particularly genes related to energy production and conversion. Notably, WP3Δhnsexhibited higher expression levels of lateral flagellar genes than WP3 and showed enhanced swarming motility and lateral flagellar production compared to those of WP3. The DNA gel mobility shift experiment showed that H-NS bound specifically to the promoter of lateral flagellar genes. Moreover, the high-affinity binding sequences of H-NS were identified by DNase I protection footprinting, and the results support the “binding and spreading” model for H-NS functioning. To our knowledge, this is the first attempt to characterize the function of the universal regulator H-NS in a deep-sea bacterium. Our data revealed that H-NS has a novel function as a repressor of the expression of genes related to the energy-consuming secondary flagellar system and to swarming motility.

Download Full-text

Acquisition of the Sda1-Encoding Bacteriophage Does Not Enhance Virulence of the Serotype M1 Streptococcus pyogenes Strain SF370

Infection and Immunity ◽

10.1128/iai.00192-13 ◽

2013 ◽

Vol 81 (6) ◽

pp. 2062-2069 ◽

Cited By ~ 16

Author(s):

Carola Venturini ◽

Cheryl-lynn Y. Ong ◽

Christine M. Gillen ◽

Nouri L. Ben-Zakour ◽

Peter G. Maamary ◽

...

Keyword(s):

Streptococcus Pyogenes ◽

Invasive Disease ◽

Mutant Form ◽

Transfer Event ◽

Content Type ◽

Lethal Infection ◽

Group A ◽

The Impact ◽

Acting In Concert

ABSTRACTThe resurgence of invasive disease caused byStreptococcus pyogenes(group AStreptococcus[GAS]) in the past 30 years has paralleled the emergence and global dissemination of the highly virulent M1T1 clone. The GAS M1T1 clone has diverged from the ancestral M1 serotype by horizontal acquisition of two unique bacteriophages, encoding the potent DNase Sda1/SdaD2 and the superantigen SpeA, respectively. The phage-encoded DNase promotes escape from neutrophil extracellular traps and is linked to enhanced virulence of the M1T1 clone. In this study, we successfully usedin vitrolysogenic conversion to transfer the Sda1-encoding phage from the M1T1 clonal strain 5448 to the nonclonal M1 isolate SF370 and determined the impact of this horizontal gene transfer event on virulence. Although Sda1 was expressed in SF370 lysogens, no capacity of the phage-converted strain to survive human neutrophil killing, switch to a hyperinvasivecovRSmutant form, or cause invasive lethal infection in a humanized plasminogen mouse model was observed. This work suggests that the hypervirulence of the M1T1 clone is due to the unique synergic effect of the M1T1 clone bacteriophage-specific virulence factor Sda1 acting in concert with the M1T1 clone-specific genetic scaffold.

Download Full-text

Regulation ofperRExpression by Iron and PerR in Campylobacter jejuni

Journal of Bacteriology ◽

10.1128/jb.05493-11 ◽

2011 ◽

Vol 193 (22) ◽

pp. 6171-6178 ◽

Cited By ~ 17

Author(s):

Minkyeong Kim ◽

Sunyoung Hwang ◽

Sangryeol Ryu ◽

Byeonghwa Jeon

Keyword(s):

Oxidative Stress ◽

Campylobacter Jejuni ◽

Regulatory Region ◽

Transcriptional Level ◽

Mobility Shift ◽

Content Type ◽

Dnase I Footprinting ◽

Transcriptional Changes ◽

Electrophoretic Mobility Shift Assays ◽

Binding Sequence

Campylobacter jejuniis a leading food-borne pathogen causing gastroenteritis in humans. Although OxyR is a widespread oxidative stress regulator in many Gram-negative bacteria,C. jejunilacks OxyR and instead possesses the metalloregulator PerR. Despite the important role played by PerR in oxidative stress defense, little is known about the factors influencingperRexpression inC. jejuni. In this study, aperRpromoter-lacZfusion assay demonstrated that iron significantly reduced the level ofperRtranscription, whereas other metal ions, such as copper, cobalt, manganese, and zinc, did not affectperRtranscription. Notably, aperRmutation substantially increased the level ofperRtranscription and intranscomplementation restored the transcriptional changes, suggestingperRis transcriptionally autoregulated inC. jejuni. In theperRmutant, iron did not repressperRtranscription, indicating the iron dependence ofperRexpression results fromperRautoregulation. Electrophoretic mobility shift assays showed that PerR binds to theperRpromoter, and DNase I footprinting assays identified a PerR binding site overlapping the −35 region of the twoperRpromoters, further supportingperRautoregulation at the transcriptional level. Alignment of the PerR binding sequence in theperRpromoter with the regulatory region of other PerR regulon genes ofC. jejunirevealed a 16-bp consensus PerR binding sequence, which shares high similarities to theBacillus subtilisPerR box. The results of this study demonstrated that PerR directly interacts with theperRpromoter and regulatesperRtranscription and thatperRautoregulation is responsible for the repression ofperRtranscription by iron inC. jejuni.

Download Full-text

Phenylacetic Acid Catabolism and Its Transcriptional Regulation in Corynebacterium glutamicum

Applied and Environmental Microbiology ◽

10.1128/aem.01588-12 ◽

2012 ◽

Vol 78 (16) ◽

pp. 5796-5804 ◽

Cited By ~ 18

Author(s):

Xi Chen ◽

Thomas A. Kohl ◽

Christian Rückert ◽

Dmitry A. Rodionov ◽

Ling-Hao Li ◽

...

Keyword(s):

Corynebacterium Glutamicum ◽

Phenylacetic Acid ◽

Specific Binding ◽

Genomic Analysis ◽

Negative Regulator ◽

Comparative Genomic ◽

Promoter Regions ◽

Mobility Shift ◽

Content Type ◽

Transcription Regulator

ABSTRACTThe industrially important organismCorynebacterium glutamicumhas been characterized in recent years for its robust ability to assimilate aromatic compounds. In this study,C. glutamicumstrain AS 1.542 was investigated for its ability to catabolize phenylacetic acid (PAA). Thepaagenes were identified; they are organized as a continuouspaagene cluster. The type strain ofC. glutamicum, ATCC 13032, is not able to catabolize PAA, but the recombinant strain ATCC 13032/pEC-K18mob2::paagained the ability to grow on PAA. ThepaaRgene, encoding a TetR family transcription regulator, was studied in detail. Disruption ofpaaRin strain AS 1.542 resulted in transcriptional increases of allpaagenes. Transcription start sites and putative promoter regions were determined. An imperfect palindromic motif (5′-ACTNACCGNNCGNNCGGTNAGT-3′; 22 bp) was identified in the upstream regions ofpaagenes. Electrophoretic mobility shift assays (EMSA) demonstrated specific binding of PaaR to this motif, and phenylacetyl coenzyme A (PA-CoA) blocked binding. It was concluded that PaaR is the negative regulator of PAA degradation and that PA-CoA is the PaaR effector. In addition, GlxR binding sites were found, and binding to GlxR was confirmed. Therefore, PAA catabolism inC. glutamicumis regulated by the pathway-specific repressor PaaR, and also likely by the global transcription regulator GlxR. By comparative genomic analysis, we reconstructed orthologous PaaR regulons in 57 species, including species ofActinobacteria,Proteobacteria, andFlavobacteria, that carry PAA utilization genes and operate by conserved binding motifs, suggesting that PaaR-like regulation might commonly exist in these bacteria.

Download Full-text

NasT-Mediated Antitermination Plays an Essential Role in the Regulation of the Assimilatory Nitrate Reductase Operon in Azotobacter vinelandii

Applied and Environmental Microbiology ◽

10.1128/aem.01720-12 ◽

2012 ◽

Vol 78 (18) ◽

pp. 6558-6567 ◽

Cited By ~ 12

Author(s):

Baomin Wang ◽

Leland S. Pierson ◽

Christopher Rensing ◽

Malkanthi K. Gunatilaka ◽

Christina Kennedy

Keyword(s):

Nitrogen Fixation ◽

Rna Binding ◽

Azotobacter Vinelandii ◽

Regulatory System ◽

Enteric Bacteria ◽

Regulatory Function ◽

Transcriptional Terminator ◽

Content Type ◽

Sequence Elements ◽

Nitrate And Nitrite

ABSTRACTAzotobacter vinelandiiis a well-studied model system for nitrogen fixation in bacteria. Regulation of nitrogen fixation inA. vinelandiiis independent of NtrB/NtrC, a conserved nitrogen regulatory system in proteobacteria. Previous work showed that anntrCmutation inA. vinelandiiresulted in a loss of induction of assimilatory nitrate and nitrite reductases encoded by thenasABoperon. In addition to NtrC, several other proteins, including NasT, a protein containing a potential RNA-binding domain ANTAR (AmiR andNasRtranscriptionantiterminationregulators), have been implicated innasABregulation. In this work, we characterize the sequence upstream ofnasAand identify several DNA sequence elements, including two potential NtrC binding sites and a putative intrinsic transcriptional terminator upstream ofnasAthat are potentially involved innasABregulation. Our analyses confirm that thenasABpromoter,PnasA, is under NtrC control. However, unlike NtrC-regulated promoters in enteric bacteria,PnasAshows high activity in the presence of ammonium; in addition, thePnasAactivity is altered in thenifAgene mutation background. We discuss the implication of these results on NtrC-mediated regulation inA. vinelandii. Our study provides direct evidence that induction ofnasABis regulated by NasT-mediated antitermination, which occurs within the leader region of the operon. The results also support the hypothesis that NasT binds the promoter proximal hairpin ofnasABfor its regulatory function, which contributes to the understanding of the regulatory mechanism of ANTAR-containing antiterminators.

Download Full-text