scholarly journals Transcriptome and Functional Analysis of the Eukaryotic-Type Serine/Threonine Kinase PknB in Staphylococcus aureus

2009 ◽  
Vol 191 (13) ◽  
pp. 4056-4069 ◽  
Author(s):  
Stefanie Donat ◽  
Karin Streker ◽  
Tanja Schirmeister ◽  
Sonja Rakette ◽  
Thilo Stehle ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Biochemical Characterization ◽  
Dependent Manner ◽  
Serine Threonine Kinase ◽  
Expression Of Genes ◽  
Biochemical Assays ◽  
Genes Encoding ◽  
Regulatory Impact ◽  
Glutamine Synthesis

ABSTRACT The function of the Staphylococcus aureus eukaryotic-like serine/threonine protein kinase PknB was investigated by performing transcriptome analysis using DNA microarray technology and biochemical assays. The transcriptional profile revealed a strong regulatory impact of PknB on the expression of genes encoding proteins which are involved in purine and pyrimidine biosynthesis, cell wall metabolism, autolysis, and glutamine synthesis. Functional activity of overexpressed and purified PknB kinase was demonstrated using the myelin basic protein as a surrogate substrate. Phosphorylation occurred in a time-dependent manner with Mn2+ as a preferred cofactor. Furthermore, biochemical characterization revealed regulation of adenylosuccinate synthase (PurA) activity by phosphorylation. Phosphorylated PurA showed a 1.8-fold decrease in enzymatic activity compared to unphosphorylated PurA. Loss of PknB led to formation of larger cell clusters, and a pknB deletion strain showed 32-fold-higher sensitivity to the cell wall-active antibiotic tunicamycin. The results of this study strongly indicate that PknB has a role in regulation of purine biosynthesis, autolysis, and central metabolic processes in S. aureus.

scholarly journals Global Regulatory Impact of ClpP Protease of Staphylococcus aureus on Regulons Involved in Virulence, Oxidative Stress Response, Autolysis, and DNA Repair

2006 ◽  
Vol 188 (16) ◽  
pp. 5783-5796 ◽  
Author(s):  
Antje Michel ◽  
Franziska Agerer ◽  
Christof R. Hauck ◽  
Mathias Herrmann ◽  
Joachim Ullrich ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Staphylococcus Aureus ◽  
Dna Repair ◽  
Stress Response ◽  
Oxidative Stress Response ◽  
Transcriptional Analysis ◽  
Expression Of Genes ◽  
Wide Range ◽  
Genes Encoding ◽  
Regulatory Impact

ABSTRACT Staphylococcus aureus is an important pathogen, causing a wide range of infections including sepsis, wound infections, pneumonia, and catheter-related infections. In several pathogens ClpP proteases were identified by in vivo expression technologies to be important for virulence. Clp proteolytic complexes are responsible for adaptation to multiple stresses by degrading accumulated and misfolded proteins. In this report clpP, encoding the proteolytic subunit of the ATP-dependent Clp protease, was deleted, and gene expression of ΔclpP was determined by global transcriptional analysis using DNA-microarray technology. The transcriptional profile reveals a strong regulatory impact of ClpP on the expression of genes encoding proteins that are involved in the pathogenicity of S. aureus and adaptation of the pathogen to several stresses. Expression of the agr system and agr-dependent extracellular virulence factors was diminished. Moreover, the loss of clpP leads to a complete transcriptional derepression of genes of the CtsR- and HrcA-controlled heat shock regulon and a partial derepression of genes involved in oxidative stress response, metal homeostasis, and SOS DNA repair controlled by PerR, Fur, MntR, and LexA. The levels of transcription of genes encoding proteins involved in adaptation to anaerobic conditions potentially regulated by an Fnr-like regulator were decreased. Furthermore, the expression of genes whose products are involved in autolysis was deregulated, leading to enhanced autolysis in the mutant. Our results indicate a strong impact of ClpP proteolytic activity on virulence, stress response, and physiology in S. aureus.

scholarly journals Variable Expressions of Staphylococcus aureus Bicomponent Leucotoxins Semiquantified by Competitive Reverse Transcription-PCR

2000 ◽  
Vol 66 (9) ◽  
pp. 3931-3938 ◽  
Author(s):  
St�phane Bronner ◽  
Patricia Stoessel ◽  
Alain Gravet ◽  
Henri Monteil ◽  
Gilles Pr�vost
Keyword(s):  
Staphylococcus Aureus ◽  
Reverse Transcription ◽  
Regulatory System ◽  
Optimization Procedure ◽  
Reverse Transcription Pcr ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Specific Mrnas ◽  
Casamino Acids ◽  
Global Regulatory System

ABSTRACT A competitive reverse transcription-PCR method was developed for the semiquantitation of the expression of genes encoding bicomponent leucotoxins of Staphylococcus aureus, e.g., Panton-Valentine leucocidin (lukPV), gamma-hemolysin (hlgA and hlgCB), and LukE-LukD (lukED). The optimization procedure included RNA preparation; reverse transcription; the use of various amounts of enzymes, antisense primer, and RNA; and the final amplification chain reaction. Reproducible results were obtained, with sensitivity for detection of cDNA within the range of 1 mRNA/104 CFU to 102 mRNA/CFU, depending on the gene. Both specific mRNAs were more significantly expressed at the late-exponential phase of growth. Expression was about 100-fold higher in yeast extract-Casamino Acids-pyruvate medium than in heart infusion medium. Expression of the widely distributed gamma-hemolysin locus in the NTCC 8178 strain was around 10-fold diminished compared with that in the ATCC 49775 strain. Because of the lower level of hlgA expression, the corresponding protein, which is generally not abundant in culture supernatant, should be investigated for its contribution to the leucotoxin-associated virulence. The agr, sar, and agr sar mutant strains revealed a great dependence with regard to leucotoxin expression on the global regulatory system inS. aureus, except that expression of hlgA was not affected in the agr mutant.

scholarly journals A genomic and proteomic analysis of surface proteins in bovine-adapted lineages of Staphylococcus aureus

2020 ◽  
Vol 2 (7A) ◽  
Author(s):  
Shauna D. Drumm ◽  
Rebecca Owens ◽  
Jennifer Mitchell ◽  
Orla M. Keane
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Surface Proteins ◽  
Host Cells ◽  
Mass Spectrometry Analysis ◽  
In Vitro Assays ◽  
Immune Recognition ◽  
Independent Manner ◽  
Genes Encoding

In Ireland, Staphylococcus aureus is the most common cause of intramammary infection (IMI) in cattle with the bovine-adapted lineages CC151 and CC97 most commonly found. Surface proteins play a major role in establishing and maintaining the infection. A previous study revealed that a strain from the CC151 lineage showed significant decay in genes encoding predicted surface proteins. Twenty-three S. aureus strains, twelve belonging to CC151 and eleven belonging to CC97, isolated from clinical IMI, were sequenced and genes encoding cell wall anchored (CWA) proteins predicted. Analysis showed that a minority of genes encoding putative CWA proteins were intact in the CC151 strains compared to CC97. Of the 26 known CWA proteins in S. aureus, the CC151 strains only encoded 10 intact genes while CC97 encoded on average 18 genes. Also within the CC97 lineage, the repertoire of genes varied depending on individual strains, with strains encoding between 17-20 intact genes. Although CC151 is reported to internalize within bovine host cells, it does so in a fibronectin-binding protein (FnBPA and FnBPB) independent manner. In-vitro assays were performed and results showed that strains from CC151, and surprisingly also CC97, weakly bound bovine fibronectin and that the FnBPs were poorly expressed in both these lineages. Mass spectrometry analysis of cell wall extracts revealed that SdrE and AdsA were the most highly expressed CWA proteins in both lineages. These results demonstrate significant differences between CC151 and CC97 in their repertoire of genes encoding CWA proteins, which may impact immune recognition of these strains and their interactions with host cells.

scholarly journals Basal anti-inflammatory responses in cultured endothelial cells exposed to two conjugated linoleic acids

2020 ◽  
Vol 79 (OCE2) ◽  
Author(s):  
Carina Valenzuela ◽  
Elizabeth Miles ◽  
Philip Calder
Keyword(s):  
Inflammatory Responses ◽  
Adhesion Assay ◽  
Dependent Manner ◽  
Relative Expression ◽  
Inflammatory Pathways ◽  
Expression Of Genes ◽  
Low Concentrations ◽  
Genes Encoding ◽  
Anti Inflammatory ◽  
Pre Treatment

AbstractConjugated linoleic acid (CLA) isomers have been shown to possess anti-atherosclerotic properties, which may be related to the downregulation of inflammatory pathways. Whether low concentrations of CLAs are able to affect basal, unstimulated endothelial cell (EC) responses is not clear. The aim of this study was to evaluate the effects of two CLAs (cis-9, trans-11 and trans-10, cis-12) on basal inflammatory responses by ECs.EA.hy926 cells (HUVEC lineage) were cultured under standard conditions and exposed to CLAs (1 and 10 μM) for 48 hours. MTT assay was performed to determine cell viability; incorporation of FA was confirmed by gas chromatography; inflammatory mediators were assessed by multiplex immunoassay; the relative expression of genes encoding transcription factors and inflammatory cytokines was assessed through real-time PCR and static adhesion assay was used to evaluate monocyte attachment to the EC monolayer.CLAs were incorporated into ECs in a dose-dependent manner. Pre-treatment with CLA9,11 (1 uM) significantly reduced unstimulated, basal concentrations of MCP-1 (p < 0.05), and CLA10,12 at 10 uM had the same effect (p < 0.05). Both CLAs at 10 uM increased the relative expression of NFκβ (p < 0.01 and p < 0.05, respectively), while decreasing the relative expression of PPARα (p < 0.0001), COX-2 (p < 0.0001) and IL-6 (p < 0.0001). In contrast, no effect was observed in the adhesion assay for either CLA.These results suggest that both CLAs at a low concentration have a neutral or modest anti-inflammatory effect in basal conditions, which may influence endothelial function and risk of vascular disease. Interestingly, at these low CLA concentrations some pro-inflammatory genes are upregulated while others are down regulated. This suggests complex effects of CLAs on inflammatory pathways.

scholarly journals The MAP kinase TVK1 regulates conidiation, hydrophobicity and the expression of genes encoding cell wall proteins in the fungus Trichoderma virens

Microbiology ◽  
2007 ◽  
Vol 153 (7) ◽  
pp. 2137-2147 ◽  
Author(s):  
Artemio Mendoza-Mendoza ◽  
Teresa. Rosales-Saavedra ◽  
Carlos. Cortés ◽  
Verónica. Castellanos-Juárez ◽  
Pedro. Martínez ◽  
...  
Keyword(s):  
Cell Wall ◽  
Map Kinase ◽  
Cell Wall Proteins ◽  
Trichoderma Virens ◽  
Expression Of Genes ◽  
Genes Encoding

scholarly journals PYED-1 Inhibits Biofilm Formation and Disrupts the Preformed Biofilm of Staphylococcus aureus

Antibiotics ◽  
2020 ◽  
Vol 9 (5) ◽  
pp. 240 ◽  
Author(s):  
Adriana Vollaro ◽  
Anna Esposito ◽  
Eliana Pia Esposito ◽  
Raffaele Zarrilli ◽  
Annalisa Guaragna ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Quorum Sensing ◽  
Biofilm Formation ◽  
Surface Proteins ◽  
Transcriptional Analysis ◽  
Capsular Polysaccharides ◽  
Dependent Manner ◽  
Chronic Infections ◽  
Concentration Dependent Manner ◽  
Expression Of Genes

Pregnadiene-11-hydroxy-16α,17α-epoxy-3,20-dione-1 (PYED-1), a heterocyclic corticosteroid derivative of deflazacort, exhibits broad-spectrum antibacterial activity against Gram-negative and Gram-positive bacteria. Here, we investigated the effect of PYED-1 on the biofilms of Staphylococcus aureus, an etiological agent of biofilm-based chronic infections such as osteomyelitis, indwelling medical device infections, periodontitis, chronic wound infections, and endocarditis. PYED-1 caused a strong reduction in biofilm formation in a concentration dependent manner. Furthermore, it was also able to completely remove the preformed biofilm. Transcriptional analysis performed on the established biofilm revealed that PYED-1 downregulates the expression of genes related to quorum sensing (agrA, RNAIII, hld, psm, and sarA), surface proteins (clfB and fnbB), secreted toxins (hla, hlb, and lukD), and capsular polysaccharides (capC). The expression of genes that encode two main global regulators, sigB and saeR, was also significantly inhibited after treatment with PYED-1. In conclusion, PYED-1 not only effectively inhibited biofilm formation, but also eradicated preformed biofilms of S. aureus, modulating the expression of genes related to quorum sensing, surface and secreted proteins, and capsular polysaccharides. These results indicated that PYED-1 may have great potential as an effective antibiofilm agent to prevent S. aureus biofilm-associated infections.

scholarly journals GABA Accumulation Causes Cell Elongation Defects and a Decrease in Expression of Genes Encoding Secreted and Cell Wall-Related Proteins in Arabidopsis thaliana

2011 ◽  
Vol 52 (5) ◽  
pp. 894-908 ◽  
Author(s):  
Hugues Renault ◽  
Abdelhak El Amrani ◽  
Ravishankar Palanivelu ◽  
Emily P. Updegraff ◽  
Agnès Yu ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Cell Wall ◽  
Cell Elongation ◽  
Expression Of Genes ◽  
Genes Encoding ◽  
Gaba Accumulation ◽  
Related Proteins

scholarly journals Modulation of Cell Wall Structure and Antimicrobial Susceptibility by a Staphylococcus aureus Eukaryote-Like Serine/Threonine Kinase and Phosphatase

2009 ◽  
Vol 77 (4) ◽  
pp. 1406-1416 ◽  
Author(s):  
Amanda M. Beltramini ◽  
Chitrangada D. Mukhopadhyay ◽  
Vijay Pancholi
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Gene Knockout ◽  
Critical Role ◽  
Cell Wall Structure ◽  
Wall Structure ◽  
Cellular Functions ◽  
Serine Threonine Kinase ◽  
Threonine Kinase ◽  
Irregular Cell

ABSTRACT It is well established that prokaryotes and eukaryotes alike utilize phosphotransfer to regulate cellular functions. One method by which this occurs is via eukaryote-like serine/threonine kinase (ESTK)- and phosphatase (ESTP)-regulated pathways. The role of these enzymes in Staphylococcus aureus has not yet been examined. This resilient organism is a common cause of hospital-acquired and community-associated infections, infecting immunocompromised and immunocompetent hosts alike. In this study, we have characterized a major functional ESTK (STK) and ESTP (STP) in S. aureus and found them to be critical modulators of cell wall structure and susceptibility to cell wall-acting β-lactam antibiotics. By utilizing gene knockout strategies, we created S. aureus N315 mutants lacking STP and/or STK. The strain lacking both STP and STK displayed notable cell division defects, including multiple and incomplete septa, bulging, and irregular cell size, as observed by transmission electron microscopy. Mutants lacking STP alone displayed thickened cell walls and increased resistance to the peptidoglycan-targeting glycylglycine endopeptidase lysostaphin, compared to the wild type. Additionally, mutant strains lacking STK or both STK and STP displayed increased sensitivity to cell wall-acting cephalosporin and carbapenem antibiotics. Together, these results indicate that S. aureus STK- and STP-mediated reversible phosphorylation reactions play a critical role in proper cell wall architecture, and thus the modulation of antimicrobial resistance, in S. aureus.

scholarly journals Nrg1 Is a Transcriptional Repressor for Glucose Repression of STA1 Gene Expression inSaccharomyces cerevisiae

1999 ◽  
Vol 19 (3) ◽  
pp. 2044-2050 ◽  
Author(s):  
Seok Hee Park ◽  
Sang Seok Koh ◽  
Jae Hwan Chun ◽  
Hye Jin Hwang ◽  
Hyen Sam Kang
Keyword(s):  
Gene Expression ◽  
Zinc Finger Protein ◽  
Glucose Repression ◽  
Transcriptional Repressor ◽  
Dependent Manner ◽  
Expression Of Genes ◽  
Dnase I Footprinting ◽  
Genes Encoding

ABSTRACT Expression of genes encoding starch-degrading enzymes is regulated by glucose repression in the yeast Saccharomyces cerevisiae. We have identified a transcriptional repressor, Nrg1, in a genetic screen designed to reveal negative factors involved in the expression of STA1, which encodes a glucoamylase. TheNRG1 gene encodes a 25-kDa C2H2zinc finger protein which specifically binds to two regions in the upstream activation sequence of the STA1 gene, as judged by gel retardation and DNase I footprinting analyses. Disruption of theNRG1 gene causes a fivefold increase in the level of theSTA1 transcript in the presence of glucose. The expression of NRG1 itself is inhibited in the absence of glucose. DNA-bound LexA-Nrg1 represses transcription of a target gene 10.7-fold in a glucose-dependent manner, and this repression is abolished in bothssn6 and tup1 mutants. Two-hybrid and glutathione S-transferase pull-down experiments show an interaction of Nrg1 with Ssn6 both in vivo and in vitro. These findings indicate that Nrg1 acts as a DNA-binding repressor and mediates glucose repression of the STA1 gene expression by recruiting the Ssn6-Tup1 complex.

scholarly journals Conserved Serine/Threonine Kinase Encoded by CBK1 Regulates Expression of Several Hypha-Associated Transcripts and Genes Encoding Cell Wall Proteins in Candida albicans

2002 ◽  
Vol 184 (7) ◽  
pp. 2058-2061 ◽  
Author(s):  
Mark D. McNemar ◽  
William A. Fonzi
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Morphological Differentiation ◽  
Growth Conditions ◽  
Cell Wall Proteins ◽  
Serine Threonine Kinase ◽  
General Role ◽  
Threonine Kinase ◽  
Genes Encoding ◽  
Opportunistic Fungal Pathogen

ABSTRACT The opportunistic fungal pathogen, Candida albicans, is reported to have several potential virulence factors. A potentially significant factor is the ability to undergo morphological transition from yeast to hypha. This alteration of form is accompanied by many changes within the cell, including alterations in gene expression and cell wall composition. We have isolated a gene that encodes a highly conserved serine/threonine kinase that appears to be involved in the regulation of proteins associated with the cell wall. We have assigned the designation CBK1 (cell wall biosynthesis kinase 1) to this gene. Mutants lacking CBK1 form large aggregates of round cells under all growth conditions and lack the ability to undergo morphological differentiation. Additionally, these mutants show an altered pattern of expression of several transcripts encoding proteins associated with the cell wall. The results suggest that the kinase encoded by CBK1 plays a general role in the maintenance and alteration of the cell wall of C. albicans in all morphologies.

Sign in / Sign up

Export Citation Format

Share Document