CelR-mediated activation of the cellobiose-utilization gene cluster in Streptococcus pneumoniae

The human pathogen Streptococcus pneumoniae harbours many genes encoding phosphotransferase systems and sugar ABC (ATP-binding cassette) transporters, including systems for the utilization of the β-glucoside sugar cellobiose. In this study, we show that the transcriptional regulator CelR, which has previously been found to be important for pneumococcal virulence, activates the expression of the cellobiose-utilization gene cluster (cel locus) of S. pneumoniae. Expression directed by the two promoters present in the cel locus was increased in the presence of cellobiose as sole carbon source in the medium, while expression decreased in the presence of glucose in the medium. Furthermore, we have predicted a 22 bp putative CelR regulatory site (5′-YTTTCCWTAWCAWTWAGGAAAA-3′) in the promoters of celA and celB, and in silico analysis showed that it is highly conserved in other pathogenic streptococci as well. Promoter truncations of celA and celB, where the half or full CelR regulatory site was deleted, confirmed that the CelR-binding site in PcelA and PcelB is functional. Transcriptome studies with the celR mutant and in silico prediction of the CelR regulatory site in the entire D39 genome sequence show that the cel locus is the only cluster of genes under the direct control of CelR. Therefore, CelR is a regulator dedicated to the cellobiose-dependent transcriptional activation of the cel locus.

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Fucose-Mediated Transcriptional Activation of the fcs Operon by FcsR in Streptococcus pneumoniae

Journal of Molecular Microbiology and Biotechnology ◽

10.1159/000377724 ◽

2015 ◽

Vol 25 (2-3) ◽

pp. 120-128 ◽

Cited By ~ 3

Author(s):

Irfan Manzoor ◽

Sulman Shafeeq ◽

Muhammad Afzal ◽

Oscar P. Kuipers

Keyword(s):

Streptococcus Pneumoniae ◽

Transcriptional Activation ◽

Promoter Region ◽

Transcriptional Regulator ◽

Transcriptional Activator ◽

Regulatory Site ◽

Rt Pcr ◽

The Impact

In this study, we explore the impact of fucose on the transcriptome of S. pneumoniae D39. The expression of various genes and operons, including the fucose uptake PTS and utilization operon (fcs operon) was altered in the presence of fucose. By means of quantitative RT-PCR and β-galactosidase analysis, we demonstrate the role of the transcriptional regulator FcsR, present upstream of the fcs operon, as a transcriptional activator of the fcs operon. We also predict a 19-bp putative FcsR regulatory site (5′-ATTTGAACATTATTCAAGT-3′) in the promoter region of the fcs operon. The functionality of this predicted FcsR regulatory site was further confirmed by promoter-truncation experiments, where deletion of half of the FscR regulatory site or full deletion led to the abolition of expression of the fcs operon.

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LacR Is a Repressor oflacABCDand LacT Is an Activator oflacTFEG, Constituting thelacGene Cluster in Streptococcus pneumoniae

Applied and Environmental Microbiology ◽

10.1128/aem.01370-14 ◽

2014 ◽

Vol 80 (17) ◽

pp. 5349-5358 ◽

Cited By ~ 18

Author(s):

Muhammad Afzal ◽

Sulman Shafeeq ◽

Oscar P. Kuipers

Keyword(s):

Streptococcus Pneumoniae ◽

Carbon Source ◽

Gene Cluster ◽

Sole Carbon Source ◽

Transcriptional Repressor ◽

Transcriptional Regulator ◽

Transcriptional Activator ◽

Content Type ◽

Elevated Expression

ABSTRACTComparison of the transcriptome ofStreptococcus pneumoniaestrain D39 grown in the presence of either lactose or galactose with that of the strain grown in the presence of glucose revealed the elevated expression of various genes and operons, including thelacgene cluster, which is organized into two operons, i.e.,lacoperon I (lacABCD) andlacoperon II (lacTFEG). Deletion of the DeoR family transcriptional regulatorlacRthat is present downstream of thelacgene cluster revealed elevated expression oflacoperon I even in the absence of lactose. This suggests a function of LacR as a transcriptional repressor oflacoperon I, which encodes enzymes involved in the phosphorylated tagatose pathway in the absence of lactose or galactose. Deletion oflacRdid not affect the expression oflacoperon II, which encodes a lactose-specific phosphotransferase. This finding was further confirmed by β-galactosidase assays with PlacA-lacZand PlacT-lacZin the presence of either lactose or glucose as the sole carbon source in the medium. This suggests the involvement of another transcriptional regulator in the regulation oflacoperon II, which is the BglG-family transcriptional antiterminator LacT. We demonstrate the role of LacT as a transcriptional activator oflacoperon II in the presence of lactose and CcpA-independent regulation of thelacgene cluster inS. pneumoniae.

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Transgene-Induced Silencing of the Zoosporogenesis-Specific NIFC Gene Cluster of Phytophthora infestans Involves Chromatin Alterations

Eukaryotic Cell ◽

10.1128/ec.00311-06 ◽

2007 ◽

Vol 6 (7) ◽

pp. 1200-1209 ◽

Cited By ~ 48

Author(s):

Howard S. Judelson ◽

Shuji Tani

Keyword(s):

Phytophthora Infestans ◽

Gene Cluster ◽

Sequence Similarity ◽

Transcriptional Regulator ◽

Hairpin Rna ◽

High Sequence Similarity ◽

Genes Encoding ◽

Cognate Gene ◽

Chromatin Packing ◽

Nif Proteins

ABSTRACT Clustered within the genome of the oomycete phytopathogen Phytophthora infestans are four genes encoding spore-specific nuclear LIM interactor-interacting factors (NIF proteins, a type of transcriptional regulator) that are moderately conserved in DNA sequence. NIFC1, NIFC2, and NIFC3 are zoosporogenesis-induced and grouped within 4 kb, and 20 kb away resides a sporulation-induced form, NIFS. To test the function of the NIFC family, plasmids expressing full-length hairpin constructs of NIFC1 or NIFC2 were stably transformed into P. infestans. This triggered silencing of the cognate gene in about one-third of transformants, and all three NIFC genes were usually cosilenced. However, NIFS escaped silencing despite its high sequence similarity to the NIFC genes. Silencing of the three NIFC genes impaired zoospore cyst germination by 60% but did not affect other aspects of the life cycle. Silencing was transcriptional based on nuclear run-on assays and associated with tighter chromatin packing based on nuclease accessibility experiments. The chromatin alterations extended a few hundred nucleotides beyond the boundaries of the transcribed region of the NIFC cluster and were not associated with increased DNA methylation. A plasmid expressing a short hairpin RNA having sequence similarity only to NIFC1 silenced both that gene and an adjacent member of the gene cluster, likely due to the expansion of a heterochromatic domain from the targeted locus. These data help illuminate the mechanism of silencing in Phytophthora and suggest that caution should be used when interpreting silencing experiments involving closely spaced genes.

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Inactivation of Transcriptional Regulator FabT Influences Colony Phase Variation of Streptococcus pneumoniae

mBio ◽

10.1128/mbio.01304-21 ◽

2021 ◽

Author(s):

Jinghui Zhang ◽

Weijie Ye ◽

Kaifeng Wu ◽

Shengnan Xiao ◽

Yuqiang Zheng ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Cell Surface ◽

Virulence Factors ◽

Capsular Polysaccharide ◽

Phase Variation ◽

Transcriptional Regulator ◽

Human Pathogen ◽

Content Type ◽

Reversible Phase

Streptococcus pneumoniae is a major human pathogen, and its virulence factors and especially the capsular polysaccharide have been extensively studied. In addition to virulence components that are present on its cell surface that directly interact with the host, S. pneumoniae undergoes a spontaneous and reversible phase variation that allows survival in different host environments.

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To have neighbour's fare: extending the molecular toolbox for Streptococcus pneumoniae

Microbiology ◽

10.1099/mic.0.28521-0 ◽

2006 ◽

Vol 152 (2) ◽

pp. 351-359 ◽

Cited By ~ 57

Author(s):

Tomas G. Kloosterman ◽

Jetta J. E. Bijlsma ◽

Jan Kok ◽

Oscar P. Kuipers

Keyword(s):

Streptococcus Pneumoniae ◽

Transcriptional Activation ◽

Expression System ◽

Inducible Promoter ◽

Defined Medium ◽

Reporter System ◽

Two Component System ◽

Over Expression ◽

Expression Studies ◽

Genes Encoding

In past years, several useful genetic tools have been developed to study the molecular biology of Streptococcus pneumoniae. In order to extend the existing spectrum of tools, advantage was taken of the toolbox originally developed for the closely related bacterium Lactococcus lactis, which was adapted for the manipulation of S. pneumoniae. The modified tools are as follows. (i) An improved nisin-inducible (over)expression system (NICE). The nisRK genes, encoding a two-component system essential for transcriptional activation in response to nisin, were integrated into the bgaA locus of S. pneumoniae D39. In this strain, D39nisRK, addition of nisin resulted in the overexpression of several genes placed under the control of the nisin-inducible promoter, while no detectable expression was observed in the absence of nisin. (ii) A lacZ reporter system. Using strain D39nisRK, which lacks endogenous β-galactosidase activity, the usefulness of the lacZ reporter vector pORI13 for the generation of chromosomal transcriptional fusions was demonstrated. In addition, the repA gene, necessary for the replication of pORI13, was introduced into the bgaA locus, thereby generating a background for plasmid-based promoter expression studies. (iii) A simplified chemically defined medium, which supports growth of all sequenced S. pneumoniae strains to a level comparable to that in complex medium. (iv) A system for the introduction of unmarked deletions and mutations into the chromosome, which is independent of the genotype of the target strain. Most of these systems were successfully applied in strains R6 and TIGR4 as well. In addition, the tools offer several improvements and advantages compared to existing ones. Thus, the molecular toolbox for S. pneumoniae has been successfully extended.

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Identification of genes encoding a novel ABC transporter in Lactobacillus delbrueckii for inulin polymers uptake

Scientific Reports ◽

10.1038/s41598-021-95356-1 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Yuji Tsujikawa ◽

Shu Ishikawa ◽

Iwao Sakane ◽

Ken-ichi Yoshida ◽

Ro Osawa

Keyword(s):

Bacillus Subtilis ◽

Carbon Source ◽

Heterologous Expression ◽

Gene Cluster ◽

Abc Transporter ◽

Transcriptome Analysis ◽

Sole Carbon Source ◽

Cultured Cells ◽

Lactobacillus Delbrueckii ◽

Genes Encoding

AbstractLactobacillus delbrueckii JCM 1002T grows on highly polymerized inulin-type fructans as its sole carbon source. When it was grown on inulin, a > 10 kb long gene cluster inuABCDEF (Ldb1381-1386) encoding a plausible ABC transporter was suggested to be induced, since a transcriptome analysis revealed that the fourth gene inuD (Ldb1384) was up-regulated most prominently. Although Bacillus subtilis 168 is originally unable to utilize inulin, it became to grow on inulin upon heterologous expression of inuABCDEF. When freshly cultured cells of the recombinant B. subtilis were then densely suspended in buffer containing inulin polymers and incubated, inulin gradually disappeared from the buffer and accumulated in the cells without being degraded, whereas levan-type fructans did not disappear. The results imply that inuABCDEF might encode a novel ABC transporter in L. delbrueckii to “monopolize” inulin polymers selectively, thereby, providing a possible advantage in competition with other concomitant inulin-utilizing bacteria.

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GalR Acts as a Transcriptional Activator of galKT in the Presence of Galactose in Streptococcus pneumoniae

Journal of Molecular Microbiology and Biotechnology ◽

10.1159/000439429 ◽

2015 ◽

Vol 25 (6) ◽

pp. 363-371 ◽

Cited By ~ 6

Author(s):

Muhammad Afzal ◽

Sulman Shafeeq ◽

Irfan Manzoor ◽

Oscar P. Kuipers

Keyword(s):

Streptococcus Pneumoniae ◽

Promoter Region ◽

Regulatory Mechanism ◽

Transcriptional Regulator ◽

Transcriptional Activator ◽

Regulatory Site ◽

Wild Type ◽

Rt Pcr ◽

Leloir Pathway

We explored the regulatory mechanism of Leloir pathway genes in Streptococcus pneumoniae D39. Here, we demonstrate that the expression of galKT is galactose dependent. By microarray analysis and quantitative RT-PCR, we further show the role of the transcriptional regulator GalR, present upstream of galKT, as a transcriptional activator of galKT in the presence of galactose. Moreover, we predict a 19-bp regulatory site (5′-GATAGTTTAGTAAAATTTT-3′) for the transcriptional regulator GalR in the promoter region of galK, which is also highly conserved in other streptococci. Growth comparison of D39 ΔgalK with the D39 wild type grown in the presence of galactose shows that galK is required for the proper growth of S. pneumoniae on galactose.

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McbG, a LysR Family Transcriptional Regulator, Activates the mcbBCDEF Gene Cluster Involved in the Upstream Pathway of Carbaryl Degradation in Pseudomonas sp. Strain XWY-1

Applied and Environmental Microbiology ◽

10.1128/aem.02970-20 ◽

2021 ◽

Vol 87 (9) ◽

Author(s):

Zhijian Ke ◽

Yidong Zhou ◽

Wankui Jiang ◽

Mingliang Zhang ◽

Hui Wang ◽

...

Keyword(s):

Binding Site ◽

Gene Cluster ◽

Transcriptional Activation ◽

Regulatory Mechanism ◽

Hydrolysis Product ◽

Transcriptional Regulator ◽

Transcriptional Regulators ◽

Palindromic Sequence ◽

Mobility Shift ◽

Lysr Family

ABSTRACT Although enzyme-encoding genes involved in the degradation of carbaryl have been reported in Pseudomonas sp. strain XWY-1, no regulator has been identified yet. In the mcbABCDEF cluster responsible for the upstream pathway of carbaryl degradation (from carbaryl to salicylate), the mcbA gene is constitutively expressed, while mcbBCDEF is induced by 1-naphthol, the hydrolysis product of carbaryl by McbA. In this study, we identified McbG, a transcriptional activator of the mcbBCDEF cluster. McbG is a 315-amino-acid protein with a molecular mass of 35.7 kDa. It belongs to the LysR family of transcriptional regulators and shows 28.48% identity to the pentachlorophenol (PCP) degradation transcriptional activation protein PcpR from Sphingobium chlorophenolicum ATCC 39723. Gene disruption and complementation studies reveal that mcbG is essential for transcription of the mcbBCDEF cluster in response to 1-naphthol in strain XWY-1. The results of the electrophoretic mobility shift assay (EMSA) and DNase I footprinting show that McbG binds to the 25-bp motif in the mcbBCDEF promoter area. The palindromic sequence TATCGATA within the motif is essential for McbG binding. The binding site is located between the –10 box and the transcription start site. In addition, McbG can repress its own transcription. The EMSA results show that a 25-bp motif in the mcbG promoter area plays an important role in McbG binding to the promoter of mcbG. This study reveals the regulatory mechanism for the upstream pathway of carbaryl degradation in strain XWY-1. The identification of McbG increases the variety of regulatory models within the LysR family of transcriptional regulators. IMPORTANCE Pseudomonas sp. strain XWY-1 is a carbaryl-degrading strain that utilizes carbaryl as the sole carbon and energy source for growth. The functional genes involved in the degradation of carbaryl have already been reported. However, the regulatory mechanism has not been investigated yet. Previous studies demonstrated that the mcbA gene, responsible for hydrolysis of carbaryl to 1-naphthol, is constitutively expressed in strain XWY-1. In this study, we identified a LysR-type transcriptional regulator, McbG, which activates the mcbBCDEF gene cluster responsible for the degradation of 1-naphthol to salicylate and represses its own transcription. The DNA binding site of McbG in the mcbBCDEF promoter area contains a palindromic sequence, which affects the binding of McbG to DNA. These findings enhance our understanding of the mechanism of microbial degradation of carbaryl.

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Two Pathway-Specific Transcriptional Regulators, PltR and PltZ, Coordinate Autoinduction of Pyoluteorin in Pseudomonas protegens Pf-5

Microorganisms ◽

10.3390/microorganisms9071489 ◽

2021 ◽

Vol 9 (7) ◽

pp. 1489

Author(s):

Qing Yan ◽

Mary Liu ◽

Teresa Kidarsa ◽

Colin P. Johnson ◽

Joyce E. Loper

Keyword(s):

Gene Cluster ◽

Transcriptional Regulator ◽

Transcriptional Regulators ◽

Soil Bacterium ◽

Antibiotic Biosynthesis ◽

Environmental Cues ◽

Alternative Mechanism ◽

Pseudomonas Spp ◽

Expression Of Genes ◽

Genes Encoding

Antibiotic biosynthesis by microorganisms is commonly regulated through autoinduction, which allows producers to quickly amplify the production of antibiotics in response to environmental cues. Antibiotic autoinduction generally involves one pathway-specific transcriptional regulator that perceives an antibiotic as a signal and then directly stimulates transcription of the antibiotic biosynthesis genes. Pyoluteorin is an autoregulated antibiotic produced by some Pseudomonas spp. including the soil bacterium Pseudomonas protegens Pf-5. In this study, we show that PltR, a known pathway-specific transcriptional activator of pyoluteorin biosynthesis genes, is necessary but not sufficient for pyoluteorin autoinduction in Pf-5. We found that pyoluteorin is perceived as an inducer by PltZ, a second pathway-specific transcriptional regulator that directly represses the expression of genes encoding a transporter in the pyoluteorin gene cluster. Mutation of pltZ abolished the autoinducing effect of pyoluteorin on the transcription of pyoluteorin biosynthesis genes. Overall, our results support an alternative mechanism of antibiotic autoinduction by which the two pathway-specific transcriptional regulators PltR and PltZ coordinate the autoinduction of pyoluteorin in Pf-5. Possible mechanisms by which PltR and PltZ mediate the autoinduction of pyoluteorin are discussed.

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D-galactose catabolism in archaea: operation of the DeLey–Doudoroff pathway in Haloferax volcanii

FEMS Microbiology Letters ◽

10.1093/femsle/fnaa029 ◽

2020 ◽

Vol 367 (1) ◽

Author(s):

Julia-Beate Tästensen ◽

Ulrike Johnsen ◽

Andreas Reinhardt ◽

Marius Ortjohann ◽

Peter Schönheit

Keyword(s):

Transcriptional Regulator ◽

High Specificity ◽

Comprehensive Analysis ◽

Haloferax Volcanii ◽

Knock Out ◽

Genes Encoding ◽

Functional Involvement ◽

Genome Analyses ◽

Galactose Dehydrogenase ◽

Substrate Binding Protein

ABSTRACT The haloarchaeon Haloferax volcanii was found to grow on D-galactose as carbon and energy source. Here we report a comprehensive analysis of D-galactose catabolism in H. volcanii. Genome analyses indicated a cluster of genes encoding putative enzymes of the DeLey–Doudoroff pathway for D-galactose degradation including galactose dehydrogenase, galactonate dehydratase, 2-keto-3-deoxygalactonate kinase and 2-keto-3-deoxy-6-phosphogalactonate (KDPGal) aldolase. The recombinant galactose dehydrogenase and galactonate dehydratase showed high specificity for D-galactose and galactonate, respectively, whereas KDPGal aldolase was promiscuous in utilizing KDPGal and also the C4 epimer 2-keto-3-deoxy-6-phosphogluconate as substrates. Growth studies with knock-out mutants indicated the functional involvement of galactose dehydrogenase, galactonate dehydratase and KDPGal aldolase in D-galactose degradation. Further, the transcriptional regulator GacR was identified, which was characterized as an activator of genes of the DeLey–Doudoroff pathway. Finally, genes were identified encoding components of an ABC transporter and a knock-out mutant of the substrate binding protein indicated the functional involvement of this transporter in D-galactose uptake. This is the first report of D-galactose degradation via the DeLey–Doudoroff pathway in the domain of archaea.

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