Involvement of the novel two-component NsrRS and LcrRS systems in distinct resistance pathways against nisin A and nukacin ISK-1 in Streptococcus mutans.
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SUBMITTER: Kawada-Matsuo M
PROVIDER: S-EPMC3719517 | biostudies-literature | 2013 Aug
REPOSITORIES: biostudies-literature
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Involvement of the novel two-component NsrRS and LcrRS systems in distinct resistance pathways against nisin A and nukacin ISK-1 in Streptococcus mutans.
Kawada-Matsuo Miki M Oogai Yuichi Y Zendo Takeshi T Nagao Junichi J Shibata Yukie Y Yamashita Yoshihisa Y Ogura Yoshitoshi Y Hayashi Tetsuya T Sonomoto Kenji K Komatsuzawa Hitoshi H
Applied and environmental microbiology 20130524 15
The novel two-component systems NsrRS and LcrRS are individually associated with resistance against the distinct lantibiotics nisin A and nukacin ISK-1 in Streptococcus mutans. NsrRS regulates the expression of NsrX, which is associated with nisin A binding, and LcrRS regulates the expression of the ABC transporter LctFEG. ...[more]
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Project description:Staphylococcus aureus uses two-component systems (TCSs) to adapt to stressful environmental conditions. To colonize a host, S. aureus must resist bacteriocins produced by commensal bacteria. In a comprehensive analysis using individual TCS inactivation mutants, the inactivation of two TCSs, graRS and braRS, significantly increased the susceptibility to the class I bacteriocins, nukacin ISK-1 and nisin A, and inactivation of vraSR slightly increased the susceptibility to nukacin ISK-1. In addition, two ABC transporters (BraAB and VraDE) regulated by BraRS and one transporter (VraFG) regulated by GraRS were associated with resistance to nukacin ISK-1 and nisin A. We investigated the role of these three TCSs of S. aureus in co-culture with S. warneri, which produces nukacin ISK-1, and Lactococcus lactis, which produces nisin A. When co-cultured with S. warneri or L. lactis, the braRS mutant showed a significant decrease in its population compared with the wild-type, whereas the graRS and vraSR mutants showed slight decreases. Expression of vraDE was elevated significantly in S. aureus co-cultured with nisin A/nukacin ISK-1-producing strains. These results suggest that three distinct TCSs are involved in the resistance to nisin A and nukacin ISK-1. Additionally, braRS and its related transporters played a central role in S. aureus survival in co-culture with the strains producing nisin A and nukacin ISK-1.
| S-EPMC3718698 | biostudies-literature
Project description:UnlabelledTwo-component systems (TCSs) are regulatory systems in bacteria that play important roles in sensing and adapting to the environment. In this study, we systematically evaluated the roles of TCSs in the susceptibility of the group A Streptococcus (GAS; Streptococcus pyogenes) SF370 strain to several types of lantibiotics. Using individual TCS deletion mutants, we found that the deletion of srtRK (spy_1081-spy_1082) in SF370 increased the susceptibility to nisin A, which is produced by Lactococcus lactis ATCC 11454, but susceptibility to other types of lantibiotics (nukacin ISK-1, produced by Staphylococcus warneri, and staphylococcin C55, produced by Staphylococcus aureus) was not altered in the TCS mutants tested. The expression of srtFEG (spy_1085 to spy_1087), which is located downstream of srtRK and is homologous to ABC transporters, was increased in response to nisin A. However, srtEFG expression was not induced by nisin A in the srtRK mutant. The inactivation of srtFEG increased the susceptibility to nisin A. These results suggest that SrtRK controls SrtFEG expression to alter the susceptibility to nisin A. Further experiments showed that SrtRK is required for coexistence with L. lactis ATCC 11454, which produces nisin A. Our results elucidate the important roles of S. pyogenes TCSs in the interactions between different bacterial species, including bacteriocin-producing bacteria.ImportanceIn this study, we focused on the association of TCSs with susceptibility to bacteriocins in S. pyogenes SF370, which has no ability to produce bacteriocins, and reported two major new findings. We demonstrated that the SrtRK TCS is related to susceptibility to nisin A by controlling the ABC transporter SrtFEG. We also showed that S. pyogenes SrtRK is important for survival when the bacteria are cocultured with nisin A-producing Lactococcus lactis This report highlights the roles of TCSs in the colocalization of bacteriocin-producing bacteria and non-bacteriocin-producing bacteria. Our findings provide new insights into the function of TCSs in S. pyogenes.
| S-EPMC5038018 | biostudies-literature
Project description:The two-component system (TCS) is a specific regulatory system in bacteria and plays an important role in sensing and adapting to the environment. In this study, we evaluated the roles of TCSs in the major cariogenic pathogen Streptococcus mutans in resistance to several types of bacteriocin. In a comprehensive analysis using individual TCS mutants, we found that two novel TCSs were associated with resistance against distinct lantibiotics nisin A (class I type A[I]) and nukacin ISK-1(class I type A[II]). One TCS, SMU.659-660 (designated as NsrRS), was related to resistance against nisin A produced by Lactococcus lactis ATCC 11454. The other TCS, SMU.1146-1145 (designated as LcrRS), was related to resistance against nukacin ISK-1 produced by Staphylococcus warneri ISK-1. NsrRS induced the expression of SMU.658 (designated as NsrX), which constitutes an operon with nsrRS, in response to nisin A. Inactivation of nsrX increased susceptibility to nisin A. Additionally, NsrX expression in S. mutans increased the binding affinity to nisin A compared to a no-expression strain. LcrRS induced the expression of SMU.1148-50 (lctFEG), which encodes an ABC transporter and is located upstream of lcrRS, in response to nukacin ISK-1. Inactivation of lctFEG significantly increased susceptibility to nukacin ISK-1. Electrophoretic mobility shift assay analysis revealed that NsrR and LcrR bound directly to regions upstream of nsrX and lctFEG, respectively. This is the first report that two distinct TCSs, NsrRS and LcrRS, are independently involved in resistance to nisin A and nukacin ISK-1 in S. mutans.
2013-12-31 | GSE44602 | GEO
Project description:The two-component system (TCS) is a specific regulatory system in bacteria and plays an important role in sensing and adapting to the environment. In this study, we evaluated the roles of TCSs in the major cariogenic pathogen Streptococcus mutans in resistance to several types of bacteriocin. In a comprehensive analysis using individual TCS mutants, we found that two novel TCSs were associated with resistance against distinct lantibiotics nisin A (class I type A[I]) and nukacin ISK-1(class I type A[II]). One TCS, SMU.659-660 (designated as NsrRS), was related to resistance against nisin A produced by Lactococcus lactis ATCC 11454. The other TCS, SMU.1146-1145 (designated as LcrRS), was related to resistance against nukacin ISK-1 produced by Staphylococcus warneri ISK-1. NsrRS induced the expression of SMU.658 (designated as NsrX), which constitutes an operon with nsrRS, in response to nisin A. Inactivation of nsrX increased susceptibility to nisin A. Additionally, NsrX expression in S. mutans increased the binding affinity to nisin A compared to a no-expression strain. LcrRS induced the expression of SMU.1148-50 (lctFEG), which encodes an ABC transporter and is located upstream of lcrRS, in response to nukacin ISK-1. Inactivation of lctFEG significantly increased susceptibility to nukacin ISK-1. Electrophoretic mobility shift assay analysis revealed that NsrR and LcrR bound directly to regions upstream of nsrX and lctFEG, respectively. This is the first report that two distinct TCSs, NsrRS and LcrRS, are independently involved in resistance to nisin A and nukacin ISK-1 in S. mutans. Total of 14 samples were analyzed. Total RNA from each test strain and control described below were labeled with Alexa Fluor® 555 and Alexa Fluor® 647, respectively, and were cohybridized on a single array. Labeling and hybridization were performed once or twice independently. UA159 wild type as control vs. UA159 with nisinA or nukacin ISK-1, UA159 wild type with nisinA or nukacin ISK-1 vs. nsrRS or lcrRS deletion mutant in UA159 with nisinA or nukacin ISK-1.
2013-12-31 | E-GEOD-44602 | biostudies-arrayexpress
Project description:Global transcriptional analysis of acid-inducible genes in Streptococcus mutans: multiple two-component systems involved in acid adaptation pH is a major environmental factor that regulates gene expression in many bacteria. Streptococcus mutans in dental biofilms is regularly exposed to cycles of acidic pH during the ingestion of fermentable dietary carbohydrates. The ability of S. mutans to tolerate low pH is crucial for its virulence and the pathogenesis in dental caries. To better understand its acid tolerance mechanisms, we used DNA microarray to perform genome-wide transcriptional analysis of S. mutans in response to acidic pH. The results showed that adaptation of S. mutans to pH 5.5 for 2 hrs induced differential expression of nearly 14% of genes in the genome, including 169 up-regulated genes and 108 down-regulated genes, largely categorized into six groups. Especially, we found that the genes encoding multiple two-component systems, including CiaHR, LevSR, LiaSR, ScnKR, HK/RR07 and ComDE, were up-regulated during acid adaptation. These findings were further confirmed by real time qRT-PCR and phenotypic assays of the gene deletion mutants. The results support that the multiple two-component systems are required for S. mutans to orchestrate its signal transduction networks for optimal adaptation to acidic pH.
2009-03-06 | GSE15125 | GEO
Project description:Global transcriptional analysis of acid-inducible genes in Streptococcus mutans: multiple two-component systems involved in acid adaptation pH is a major environmental factor that regulates gene expression in many bacteria. Streptococcus mutans in dental biofilms is regularly exposed to cycles of acidic pH during the ingestion of fermentable dietary carbohydrates. The ability of S. mutans to tolerate low pH is crucial for its virulence and the pathogenesis in dental caries. To better understand its acid tolerance mechanisms, we used DNA microarray to perform genome-wide transcriptional analysis of S. mutans in response to acidic pH. The results showed that adaptation of S. mutans to pH 5.5 for 2 hrs induced differential expression of nearly 14% of genes in the genome, including 169 up-regulated genes and 108 down-regulated genes, largely categorized into six groups. Especially, we found that the genes encoding multiple two-component systems, including CiaHR, LevSR, LiaSR, ScnKR, HK/RR07 and ComDE, were up-regulated during acid adaptation. These findings were further confirmed by real time qRT-PCR and phenotypic assays of the gene deletion mutants. The results support that the multiple two-component systems are required for S. mutans to orchestrate its signal transduction networks for optimal adaptation to acidic pH. Total RNAs were isolated from S. mutans UA159 cells (0.6 at OD600) grown in a TYG broth (3% tryptone, 0.3% yeast extract and 20 mM glucose) at either pH 5.5 or pH 7.5. The RNAs were treated with RNase-free DNase 1 and purified by Qiagen RNeasy mini columns. The purified RNAs were used to generate cDNA probes by an indirect labeling method based on the protocol from TIGR. The cDNAs were coupled with AlexaFluor 555 or AlexaFluor 647 (Invitrogen). The labeled cDNA probes from three different cultures of UA159 were hybridized to the S. mutans microarray slides obtained from PFGRC (http://pfgrc.tigr.org). Array hybridization was conducted using a protocol from the PFGRC with minor modification. After hybridization, washes and dried, the array slides were scanned by ScanArray 5000XL Reader (Perkin Elmer, Boston, MA). After the array slides were scanned, the resulting images were loaded into TIGR Spotfinder software (http://www.tigr.org/software/) and overlaid. A spot grid was created according to TIGR specifications and manually adjusted to fit all spots within the grid, and the intensity values of each spot were determined. Signal intensities of individual channels from an array slide were averaged and normalized using an array data analysis software (MIDAS) by using LOWESS and iterative log mean centering with default settings, followed by in-slide replicate analysis. A t-test was used to determine the consistency of ratios across replicate hybridizations. Only genes whose ratios were ⥠2-fold changes (either increase or decrease) with 99% confidence interval (P ⤠0.01) were considered statistically significant.
2010-05-22 | E-GEOD-15125 | biostudies-arrayexpress
Project description:Streptococcus mutans is a cariogenic bacterium that produces a variety of bacteriocins and retains resistance to these bacteriocins. In this study, we investigated the susceptibility of 127 S. mutans strains to nukacins produced by Staphylococcus spp., which are commensal bacteria in humans. We detected diverse susceptibilities among strains. Nineteen strains had a disrupted LctF (type I), which is responsible for nukacin susceptibility, whereas the remaining 108 strains had an intact LctF (type II) and displayed resistance to nukacins. However, the type I strains still showed resistance to nukacins to some extent. Interestingly, 18/19 (94.7%) type I strains carried a mukA-T locus, which is related to the synthesis of mutacin K8, and mukFEG, an ABC transporter. In contrast, among type II strains, only 6/108 strains (5.6%) had both the mukA-T locus and mukFEG, 19/108 strains (17.6%) carried only mukFEG, and 83/108 strains (76.9%) harbored neither mukA-T nor mukFEG. We also found that MukF had two variants: 305 amino acids (type α) and 302 amino acids (type β). All type I strains showed a type α (MukFα), whereas most type II strains with mukFEG (22/25 strains) had a type β (MukFβ). Then, we constructed a mukFEG-deletion mutant complemented with MukFαEG or MukFβEG and found that only MukFαEG was involved in nukacin resistance. The nukacin resistance capability of type II-LctFEG was stronger than that of MukFαEG. In conclusion, we identified a novel nukacin resistance factor, MukFEG, and either LctFEG or MukFEG was active in most strains via genetic polymorphisms depending on mukA-T genes.ImportanceStreptococcus mutans is an important pathogenic bacterium not only for dental caries but also for systemic diseases. S. mutans is known to produce a variety of bacteriocins and to retain resistance these bacteriocins. In this study, two ABC transporters, LctFEG and MukFEG, were implicated in nukacin resistance and each ABC transporter has two subtypes, active and inactive. Of the two ABC transporters, only one ABC transporter was always resistant, while the other ABC transporter was inactivated by genetic mutation. Interestingly, this phenomenon was defined by the presence or absence of the mutacin K8 synthesis gene region, one of the bacteriocins of S. mutans. This suggests that the resistance acquisition is tightly controlled in each strain. This study provides important evidence that the insertion of bacteriocin synthesis genes is involved in the induction of genetic polymorphisms and suggests that bacteriocin synthesis genes may play an important role in bacterial evolution.
| S-EPMC10952377 | biostudies-literature
Project description:The principal etiological agent of human dental caries, Streptococcus mutans is a multi-virulent pathogen that can transform commensal oral microbial community to plaque biofilms. Major virulence factors that are associated with the cariogenicity of S. mutans include adhesion, acidogenicity and acidurity. All these pathogenic traits coordinate and alter the dental plaque ecology which provide room for interaction with other similar acidogenic and aciduric bacteria. This cariogenic flora increases the possibility of enamel demineralization which headway to caries development. The present study was aimed at evaluating the antimicrobial and antiinfective potential of a lichen secondary metabolite usnic acid (UA) against S. mutans. Minimum inhibitory concentration (MIC), Minimum bactericidal concentration (MBC) and growth kinetics were evaluated to determine the antimicrobial potential of UA against S. mutans. UA at 5 µg mL-1 and 10 µg mL-1 concentration were considered as MIC and MBC respectively. Effect on biofilm formation was microscopically assessed and found to be reduced in a concentration dependent manner. Gene expression of gtfB, gtfC, gtfD, vicR, ComDE and smu0630 was found to be downregulated upon treatment with sub-MIC of UA. Acidogenicity, acidurity, eDNA synthesis and response to oxidative stress were found to be attenuated by the influence of UA. It was also demonstrated to act on preformed mature biofilm of S. mutans. Moreover, UA was shown to possess very low frequency to acquire spontaneous resistance development in S. mutans. Besides, no morphological aberrations or toxic effect was instigated by UA in the human buccal epithelial cells as well as to the oral commensals. Altogether, these results demonstrate the therapeutic potential of usnic acid in the treatment of S. mutans infection.
| S-EPMC7809355 | biostudies-literature
Project description:The LiaSR two-component signal transduction system regulates cellular responses to several environmental stresses, including those that induce cell envelope damages. Downstream regulons of the LiaSR system have been implicated in tolerance to acid, antibiotics and detergents. In the dental pathogen Streptococcus mutans, the LiaSR system is necessary for tolerance against acid, antibiotics, and cell wall damaging stresses during growth in the oral cavity. To understand the molecular mechanisms by which LiaSR regulates gene expression, we created a mutant LiaR in which the conserved aspartic acid residue (the phosphorylation site), was changed to alanine residue (D58A). As expected, the LiaR-D58A variant was unable to acquire the phosphate group and bind to target promoters. We also noted that the predicted LiaR-binding motif upstream of the lia operon does not appear to be well conserved. Consistent with this observation, we found that LiaR was unable to bind to the promoter region of lia; however, we showed that LiaR was able to bind to the promoters of SMU.753, SMU.2084 and SMU.1727. Based on sequence analysis and DNA binding studies we proposed a new 25-bp conserved motif essential for LiaR binding. Introducing alterations at fully conserved positions in the 25-bp motif affected LiaR binding, and the binding was dependent on the combination of positions that were altered. By scanning the S. mutans genome for the occurrence of the newly defined LiaR binding motif, we identified the promoter of hrcA (encoding a key regulator of the heat shock response) that contains a LiaR binding motif, and we showed that hrcA is negatively regulated by the LiaSR system. Taken together our results suggest a putative role of the LiaSR system in heat shock responses of S. mutans.
| S-EPMC4447274 | biostudies-literature
Project description:The gram-positive bacterium Streptococcus mutans is the primary causative agent in the formation of dental caries in humans. The ability of S. mutans to adapt and to thrive in the hostile environment of the oral cavity suggests that this cariogenic pathogen is capable of sensing and responding to different environmental stimuli. This prompted us to investigate the role of two-component signal transduction systems (TCS), particularly the sensor kinases, in response to environmental stresses. Analysis of the annotated genome sequence of S. mutans indicates the presence of 13 putative TCS. Further bioinformatics analysis in our laboratory has identified an additional TCS in the genome of S. mutans. We verified the presence of the 14 sensor kinases by using PCR and Southern hybridization in 13 different S. mutans strains and found that not all of the sensor kinases are encoded by each strain. To determine the potential role of each TCS in the stress tolerance of S. mutans UA159, insertion mutations were introduced into the genes encoding the individual sensor kinases. We were successful in inactivating all of the sensor kinases, indicating that none of the TCS are essential for the viability of S. mutans. The mutant S. mutans strains were assessed for their ability to withstand various stresses, including osmotic, thermal, oxidative, and antibiotic stress, as well as the capacity to produce mutacin. We identified three sensor kinases, Smu486, Smu1128, and Smu1516, which play significant roles in stress tolerance of S. mutans strain UA159.
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