katA Gene Research Articles

CzcR-dependent reduction of catalase gene expression and induction of catalase activity in Pseudomonas aeruginosa during zinc excess

BackgroundPseudomonas aeruginosa is able to survive, grow, and cause severe infections at different sites throughout the human body owing to its ability to sense diverse signals and precisely modulate target gene expression using its abundant signaling systems. Release of zinc (Zn) and hydrogen peroxide (H2O2) within the phagocyte are two major host strategies to defend against bacterial infections. It was previously shown that the response regulator CzcR controls global gene expression including catalase genes during Zn excess, but regulatory mechanisms of catalase gene expression and the role of CzcR in H2O2 tolerance remain unclear.ResultsIn the study, comparative transcriptome analysis comprehensively described the CzcR-dependent and -independent gene regulatory pattern in P. aeruginosa during Zn excess, which revealed the counteractive co-regulation of two key H2O2-detoxifying catalase genes katA and katB through CzcR-dependent and -independent pathways in response to Zn excess. Protein-DNA interaction assay demonstrated that CzcR negatively regulates the expression of catalase genes katA and katB by directly binding to their promoters. While interestingly, we further showed that CzcR positively regulates H2O2 tolerance by inducing the catalase activity during Zn excess.ConclusionThis study reported the opposite functions of CzcR in negatively regulating the expression of catalase genes katA and katB but in positively regulating the activity of catalase and H2O2 tolerance during Zn excess in P. aeruginosa.

The enhanced antibacterial effect and mechanism of PostbioYDFF-3 against Pseudomonas putida NBRC 14164 and Acinetobacter johnsonii ANC 3681

The inhibitory mechanism of postbioYDFF-3 (postbiotics) on Pseudomonas putida NBRC 14164 (P. putida NBRC 14164) and Acinetobacter johnsonii ANC 3681 (A. johnsonii ANC 3681) in refrigerated fish fillet was investigated. At the cellular level, the minimum inhibitory concentrations of P. putida NBRC 14164 and A. johnsonii ANC 3681 treated with postbiotics were 3.9063 μL/mL and 15.625 μL/mL, respectively. The postbiotics effectively inhibited biofilm formation, promoted cell DNA leakage and increased cellular membrane permeability. Furthermore, the transcriptomic results showed that the pathways of transmembrane transport, energy metabolism, DNA damage repair, nucleotide binding and oxidation-reduction (REDOX) were significantly down-regulated. In particular, PhnA and A1_GM000335 genes were up-regulated by 9.71 and 7.16 times respectively, which was the key to stress response in the nucleotide binding pathway of A. johnsonii ANC 3681. PP4_RS22590 and katE genes were down-regulated by 28.64 and 11.63 times respectively, which caused the REDOX inhibition of P. putida NBRC 14164. The transcriptomic results were verified by quantitative real-time PCR (qRT-PCR). This study provided a new theoretical basis for the inhibition of postbiotics against P. putida NBRC 14164 and A. johnsonii ANC 3681.

Metabolic engineering of Escherichia coli for the production of 5-aminolevulinic acid based on combined metabolic pathway modification and reporter-guided mutant selection (RGMS)

Background5-Aminolevulinic acid (ALA) recently received much attention due to its potential application in many fields such as medicine, nutrition and agriculture. Metabolic engineering is an efficient strategy to improve microbial production of 5-ALA.ResultsIn this study, an ALA production strain of Escherichia coli was constructed by rational metabolic engineering and stepwise improvement. A metabolic strategy to produce ALA directly from glucose in this recombinant E. coli via both C4 and C5 pathways was applied herein. The expression of a modified hemARS gene and rational metabolic engineering by gene knockouts significantly improved ALA production from 765.9 to 2056.1 mg/L. Next, we tried to improve ALA production by RGMS-directed evolution of eamA gene. After RGMS, the ALA yield of strain A2-ASK reached 2471.3 mg/L in flask. Then, we aimed to improve the oxidation resistance of cells by overexpressing sodB and katE genes and ALA yield reached 2703.8 mg/L. A final attempt is to replace original promoter of hemB gene in genome with a weaker one to decrease its expression. After 24 h cultivation, a high ALA yield of 19.02 g/L was achieved by 108-ASK in a 5 L fermenter.ConclusionsThese results suggested that an industrially competitive strain can be efficiently developed by metabolic engineering based on combined rational modification and optimization of gene expression.

Inter-species Transcriptomic Analysis Reveals a Constitutive Adaptation Against Oxidative Stress for the Highly Virulent Leptospira Species.

Transcriptomic analyses across large scales of evolutionary distance have great potential to shed light on regulatory evolution but are complicated by difficulties in establishing orthology and limited availability of accessible software. We introduce here a method and a graphical user interface wrapper, called Annotator-RNAtor, for performing interspecies transcriptomic analysis and studying intragenus evolution. The pipeline uses third-party software to infer homologous genes in various species and highlight differences in the expression of the core-genes. To illustrate the methodology and demonstrate its usefulness, we focus on the emergence of the highly virulent Leptospira subclade known as P1+, which includes the causative agents of leptospirosis. Here, we expand on the genomic study through the comparison of transcriptomes between species from P1+ and their related P1- counterparts (low-virulent pathogens). In doing so, we shed light on differentially expressed pathways and focused on describing a specific example of adaptation based on a differential expression of PerRA-controlled genes. We showed that P1+ species exhibit higher expression of the katE gene, a well-known virulence determinant in pathogenic Leptospira species correlated with greater tolerance to peroxide. Switching PerRA alleles between P1+ and P1- species demonstrated that the lower repression of katE and greater tolerance to peroxide in P1+ species was solely controlled by PerRA and partly caused by a PerRA amino-acid permutation. Overall, these results demonstrate the strategic fit of the methodology and its ability to decipher adaptive transcriptomic changes, not observable by comparative genome analysis, that may have been implicated in the emergence of these pathogens.

Molecular epidemiology, genetic diversity, antibiotic resistance and pathogenicity of Stenotrophomonas maltophilia complex from bacteremia patients in a tertiary hospital in China for nine years.

The Stenotrophomonas maltophilia complex (Smc) has emerged as a significant nosocomial pathogen contributing to increased mortality rates, particularly in case of bloodstream infections. This study employed whole-genome sequencing (WGS) to assess the genetic diversity, antimicrobial resistance profiles, molecular epidemiology and frequencies of virulence genes among 55 S. maltophilia isolates obtained from bacteremic cases over a 9-year period. Based on the threshold of 95% average nucleotide identity (ANI) and 70% digital DNA-DNA hybridization (dDDH) for genospecies delineation, we classified 37 isolates into 6 known species, all belonging to the Smc. The remaining 18 isolates sequenced in this study were assigned to 6 new genomospecies. Among the 55 isolates, we identified 44 different sequence types (STs), comprising 22 known and 22 novel allele combinations. The resistance rate of Smc against trimethoprim-sulfamethoxazole (TMP/SMX) was found to be 3.6%, with the sul1 and class one integron integrase genes (intI) detected in these isolates. All Smc isolates were susceptible to minocycline. Furthermore, all Smc strains harbored the motA, pilU, smf-1 and Stmpr2 genes. Genomospecies 1 (100%, n = 9), Stenotrophomonas maltophilia (84.21%, n = 19) and Stenotrophomonas sepilia (71.43%, n = 7) demonstrated a higher percentage of the afaD gene, which was also associated with a higher separation rate. In addition to motA, pilU, smf-1, and Stmpr2 genes, all S. maltophilia strains (100%) contained entA, gspD, KatA, and stmPr1 genes, while all genomospecies 1 strains (100%) contained afaD, entA, gspD, and KatA genes. Our study highlights the genetic diversity among Smc isolates from patients with bacteremia, revealing 22 novel ST types, 58 new alleles and 6 new genomospecies. S. maltophilia and S. pavanii were found to carry more virulence factors, emphasizing the importance of accurate strain identification. Minocycline emerged as a promising alternative antibiotic for patients who were resistant to TMP/SMX.

Changes in Activity of Antioxidant Systems of <i>Escherichia coli</i> under Phosphate Starvation

Changes in the activity of antioxidant systems in Escherichia coli during phosphate starvation were studied. It was shown that starvation was accompanied by a decrease in the intensity of respiration, an increase in the rate of superoxide production, and a decrease in the level of ATP. Simultaneously, there was a decrease in H2O2 in the medium and a significant increase in the expression of the katG and katE genes encoding the HPI and HPII catalases, respectively. At the same time, there was no drop in the membrane potential, which may indicate the retention of normal membrane activity in starving cells. It has been shown for the first time that the transition of E. coli to phosphate starvation is accompanied by significant changes in the status of glutathione. The most important of them are associated with a decrease in the level of glutathione reductive form (GSH) in the medium (GSHout) and with a simultaneous increase in its content in the cytoplasm (GSHin), as well as a shift in the GSHin to oxidized glutathione form (GSSGin) ratio towards reductive values, and GSHout/GSSGout towards oxidative values. Among the mutants used in the work, the double mutant gor trxB, deficient in the synthesis of glutathione reductase and thioredoxin reductase, showed the most pronounced distinctive features. Compared to the parental strain, this mutant showed a multiple higher expression of katG::lacZ, the highest level of oxidized intra- and extracellular glutathione, and, accordingly, the lowest GSH/GSSG ratio in both compartments. In general, the data obtained indicate that during phosphate starvation the interaction of the glutathione redox-system and regulons that control protection against reactive oxygen species creates conditions that allow maintaining the concentration of ROS below the toxic level. As a result, phosphate-starved E. coli cells can maintain a high viability for a long time that allows them quickly to resume growth after the addition of phosphate.

Functionality, characterization and DEGs contribution by engineering isolate Pseudomonas P1 to elucidate the regulation mechanisms of p-chlorophenol-4-Chloroaniline bioremediation

Engineered bacteria are one of the greenest and most profitable biodegradation methods for removing noxious chlorophenols and chloramines from the environment. This study aims to acquire an engineering isolate Pseudomonas P1 that can effectively degrade 4-Chloroaniline (4-CA) and p-Chlorophenol (4-CP) simultaneously, and reveal potential differences in core gene expression and metabolic pathways in practical applications. A protein interaction network diagram was drawn using networkX under Python to count various topological properties of genes interaction network based on whole genome sequencing to reveal the interaction between proteins involved in organochlorine degradation. The core genes expression levels during aerobic degradation of the aforementioned compounds by Pseudomonas P1, which the most excellent degradation specifications at 30 °C, neutral pH and 100 mg/L, was analyzed based on transcriptome sequencing. Importantly, two major pathways exist in Pseudomonas P1 based on transcriptome sequencing during aerobic degradation of the aforementioned compounds. 4-chlorocatechol (4-CC) pathway of 4-CP degradation by multicomponent phenol hydroxylase (DmpONMLKP) and catechol 2,3-dioxygenase (DmpB), which expression levels were notably up-regulated 1.1–3.25 folds and DmpB was the most significant increased. GSH-dependent dehalogenation pathway including GSR, acpD, ahpF, yghU and katE genes to degrade 4-CA, which expression levels were significantly up-regulated 1.0–3.5 folds. The core genes expression changes were further verified by RTq-PCR. Collectively, our results provided an engineered bacteria that can efficiently mineralize 4-CA and 4-CP simultaneously, and clarified that it has two metabolic pathways regulate biodegradation of 4-CA and 4-CP, which can be practiced for in-site bioremediation of chlorophenols and chloramines co-contaminated soil.

ArcR contributes to tolerance to fluoroquinolone antibiotics by regulating katA in Staphylococcus aureus.

Staphylococcus aureus is an opportunistic pathogen that shows a unique ability to quickly respond to a variety of antibiotics. The Crp/Fnr family transcriptional regulator ArcR controls expression of arginine deiminase pathway genes arcABDC, which enable the utilization of arginine as an energy source for cell growth under anaerobic conditions. However, ArcR shares low overall similarity with other Crp/Fnr family proteins, suggesting that they differ in the response to environmental stress. In this study, MIC and survival assays were performed to determine the role of ArcR in antibiotic resistance and tolerance. The results showed that deletion of arcR reduced tolerance of S.aureus to fluoroquinolone antibiotics, mainly through a defect in the response to oxidative stress. In ΔarcR mutant, the expression of the major catalase gene katA was downregulated, and katA overexpression restored bacterial resistance to oxidative stress and antibiotics. We showed that ArcR directly regulated katA transcription by binding to the promoter region of katA. Therefore, our results revealed the contribution of ArcR in bacterial tolerance to oxidative stress and subsequently to fluoroquinolones antibiotics. This study added our understanding on the role of Crp/Fnr family in bacterial susceptibility to antibiotics.

The enigmatic physiological roles of AhpCF, Gpx, Npr and Kat in peroxide stress response of Enterococcus faecium

In this work, the physiological roles of the primary peroxide scavenging activities of Enterococcus faecium AUS0004 strain were analysed. This healthcare-associated pathogen harbours genes encoding putative NADH peroxidase (Npr), alkyl hydroperoxide reductase (AhpCF), glutathione peroxidase (Gpx) and manganese-dependent catalase (Mn-Kat). Gene expression analyses showed that npr and kat genes are especially and significantly induced in cells treated with hydrogen peroxide (H2O2) and cumene hydroperoxide (CuOOH), which suggested an important function of these enzymes to protect E. faecium against peroxide stress. Mutants affected in one or several predicted anti-oxidative activities mentioned above showed that neither the peroxidases nor the catalase are implicated in the defence against peroxide challenges. However, our investigations allowed us to show that Npr is responsible for the degradation of approximately 45% of metabolically derived H2O2 which avoids accumulation of the peroxide to lethal concentrations.

The Pseudomonas aeruginosa DksA1 protein is involved in H2O2 tolerance and within-macrophages survival and can be replaced by DksA2

In Gram-negative pathogens, the stringent response regulator DksA controls the expression of hundreds of genes, including virulence-related genes. Interestingly, Pseudomonas aeruginosa has two functional DksA paralogs: DksA1 is constitutively expressed and has a zinc-finger motif, while DksA2 is expressed only under zinc starvation conditions and does not contain zinc. DksA1 stimulates the production of virulence factors in vitro and is required for full pathogenicity in vivo. DksA2 can replace these DksA1 functions. Here, the role of dksA paralogs in P. aeruginosa tolerance to H2O2-induced oxidative stress has been investigated. The P. aeruginosa dksA1 dksA2 mutant showed impaired H2O2 tolerance in planktonic and biofilm-growing cultures and increased susceptibility to macrophages-mediated killing compared to the wild type. Complementation with either dksA1 or dksA2 genes restored the wild type phenotypes. The DksA-dependent tolerance to oxidative stress involves, at least in part, the positive transcriptional control of both katA and katE catalase-encoding genes. These data support the hypothesis that DksA1 and DksA2 are eco-paralogs with indistinguishable function but optimal activity under different environmental conditions, and highlight their mutual contribution to P. aeruginosa virulence.

Genomic Analysis of a Highly Virulent NDM-1-Producing Escherichia coli ST162 Infecting a Pygmy Sperm Whale (Kogia breviceps) in South America.

Carbapenemase-producing Enterobacterales are rapidly spreading and adapting to different environments beyond hospital settings. During COVID-19 lockdown, a carbapenem-resistant NDM-1-positive Escherichia coli isolate (BA01 strain) was recovered from a pygmy sperm whale (Kogia breviceps), which was found stranded on the southern coast of Brazil. BA01 strain belonged to the global sequence type (ST) 162 and carried the blaNDM–1, besides other medically important antimicrobial resistance genes. Additionally, genes associated with resistance to heavy metals, biocides, and glyphosate were also detected. Halophilic behavior (tolerance to > 10% NaCl) of BA01 strain was confirmed by tolerance tests of NaCl minimal inhibitory concentration, whereas halotolerance associated genes katE and nhaA, which encodes for catalase and Na+/H+ antiporter cytoplasmic membrane, respectively, were in silico confirmed. Phylogenomics clustered BA01 with poultry- and human-associated ST162 lineages circulating in European and Asian countries. Important virulence genes, including the astA (a gene encoding an enterotoxin associated with human and animal infections) were detected, whereas in vivo experiments using the Galleria mellonella infection model confirmed the virulent behavior of the BA01 strain. WHO critical priority carbapenemase-producing pathogens in coastal water are an emerging threat that deserves the urgent need to assess the role of the aquatic environment in its global epidemiology.

Sinorhizobium melilotiohrR genes affect symbiotic performance with alfalfa (Medicago sativa).

Sinorhizobium meliloti infects the host plant alfalfa to induce formation of nitrogen-fixation root nodules, which inevitably elicit reactive oxygen species (ROS) bursts and organic peroxide generation. The MarR family regulator OhrR regulates the expression of chloroperoxidase and organic hydrogen resistance protein, which scavenge organic peroxides in free-living S. meliloti cells. The single mutant of ohrR genes SMc01945 (ohrR1) and SMc00098 (ohrR2) lacked symbiotic phenotypes. In this work, we identified the novel ohrR gene SMa2020 (ohrR3) and determined that ohrR genes are important for rhizobial infection, nodulation and nitrogen fixation with alfalfa. By analysing the phenotypes of the single, double and triple deletion mutants of ohrR genes, we demonstrate that ohrR1 and ohrR3 slightly affect rhizobial growth, but ohrR2 and ohrR3 influence cellular resistance to the organic peroxide, tert-butyl hydroperoxide. Deletion of ohrR1 and ohrR3 negatively affected infection thread formation and nodulation, and consequently, plant growth. Correspondingly, the expression of the ROS detoxification genes katA and sodB as well as that of the nitrogenase gene nifH was downregulated in bacteroids of the double and triple deletion mutants, which may underlie the symbiotic defects of these mutants. These findings demonstrate that OhrR proteins play a role in the S. meliloti-alfalfa symbiosis.

The OxyR and SoxR transcriptional regulators are involved in a broad oxidative stress response in Paraburkholderia xenovorans LB400

BackgroundAerobic metabolism generates reactive oxygen species that may cause critical harm to the cell. The aim of this study is the characterization of the stress responses in the model aromatic-degrading bacterium Paraburkholderia xenovorans LB400 to the oxidizing agents paraquat and H2O2.MethodsAntioxidant genes were identified by bioinformatic methods in the genome of P. xenovorans LB400, and the phylogeny of its OxyR and SoxR transcriptional regulators were studied. Functionality of the transcriptional regulators from strain LB400 was assessed by complementation with LB400 SoxR of null mutant P. aeruginosa ΔsoxR, and the construction of P. xenovorans pIZoxyR that overexpresses OxyR. The effects of oxidizing agents on P. xenovorans were studied measuring bacterial susceptibility, survival and ROS formation after exposure to paraquat and H2O2. The effects of these oxidants on gene expression (qRT-PCR) and the proteome (LC–MS/MS) were quantified.ResultsP. xenovorans LB400 possesses a wide repertoire of genes for the antioxidant defense including the oxyR, ahpC, ahpF, kat, trxB, dpsA and gorA genes, whose orthologous genes are regulated by the transcriptional regulator OxyR in E. coli. The LB400 genome also harbors the soxR, fumC, acnA, sodB, fpr and fldX genes, whose orthologous genes are regulated by the transcriptional regulator SoxR in E. coli. The functionality of the LB400 soxR gene was confirmed by complementation of null mutant P. aeruginosa ΔsoxR. Growth, susceptibility, and ROS formation assays revealed that LB400 cells were more susceptible to paraquat than H2O2. Transcriptional analyses indicated the upregulation of the oxyR, ahpC1, katE and ohrB genes in LB400 cells after exposure to H2O2, whereas the oxyR, fumC, ahpC1, sodB1 and ohrB genes were induced in presence of paraquat. Proteome analysis revealed that paraquat induced the oxidative stress response proteins AhpCF and DpsA, the universal stress protein UspA and the RNA chaperone CspA. Both oxidizing agents induced the Ohr protein, which is involved in organic peroxide resistance. Notably, the overexpression of the LB400 oxyR gene in P. xenovorans significantly decreased the ROS formation and the susceptibility to paraquat, suggesting a broad OxyR-regulated antioxidant response.ConclusionsThis study showed that P. xenovorans LB400 possess a broad range oxidative stress response, which explain the high resistance of this strain to the oxidizing compounds paraquat and H2O2.

Unmarked Gene Editing in Clavibacter michiganensis Using CRISPR/Cas9 and 5-Fluorocytosine Counterselection.

Plant-pathogenic bacteria in the genus Clavibacter are important quarantine species that cause considerable economic loss worldwide. The development of effective gene editing techniques and additional selectable markers is essential to expedite gene functional analysis in this important Gram-positive genus. The current study details a highly efficient unmarked CRISPR/Cas9-mediated gene editing system in Clavibacter michiganensis, which couples the expression of cas9 and single-guide RNA with homology-directed repair templates and the negative selectable marker codA::upp within a single plasmid. Initial experiments indicated that CRISPR/Cas9-mediated transformation could be utilized for both site-directed mutagenesis, in which an A to G point mutation was introduced at the 128th nucleotide of the C. michiganensis rpsL gene to generate a streptomycin-resistant mutant, and complete gene knockout, in which the deletion of the C. michiganensis celA or katA genes resulted in transformants that lacked cellulase and catalase activity, respectively. In subsequent experiments, the introduction of the codA::upp cassette into the transformation vector facilitated the counterselection of unmarked transformants by incubation in the absence of the selective antibiotic, followed by plating on M9 agar containing 5-fluorocytosine at 100 μg/ml, in which an unmarked katA mutant lacking the transformation vector was recovered. Compared with conventional homologous recombination, the unmarked CRISPR/Cas9-mediated system was more useful and convenient because it allowed the template plasmid to be reused repeatedly to facilitate the editing of multiple genes, which constitutes a major advancement that could revolutionize research into C. michiganensis and other Clavibacter spp.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Drought‐induced increase in catalase activity improves cotton yield when grown under water‐limiting field conditions

AbstractPlants experiencing abiotic stress can produce excessive reactive oxygen species (ROS), ultimately impacting the crop growth and yield. Hydrogen peroxide H2O2 is a relatively stable active ROS and is mainly scavenged by catalase. In this study, the catalase‐encoding E. coli KatE gene was transferred to cotton plants by Agrobacterium‐mediated transformation. From almost 100 independent transgenic lines, three transgenic lines were identified that did not display a discernible growth and yield phenotype when grown under well‐watered field conditions, but were tolerant to drought under field conditions, resulting in a ~ 25% higher cotton‐seed yield. The three lines were analysed under controlled laboratory conditions to explain the drought‐dependent phenotype: the transgenic lines displayed drought‐inducible EcKatE expression and total catalase activity was only higher than that of the wild type during growth under drought or salinity stress. Furthermore, drought‐induced ectopic EcKatE expression increased total SOD and POD activities, decreased H2O2 levels and improved photosystem II performance and relative water content. The results show that ectopic EcKatE expression displays an advantage during drought stress, without negatively affecting yield during well‐watered field conditions. EcKatE overexpression confers hyper‐resistance to drought in cotton and provides improved cotton‐seed yield under water‐limiting field conditions. And the EcKatE gene might also be a promising drought‐resistant gene for improving other crop varieties.

A peroxisomal β-oxidative pathway contributes to the formation of C6-C1 aromatic volatiles in poplar.

Benzenoids (C6–C1 aromatic compounds) play important roles in plant defense and are often produced upon herbivory. Black cottonwood (Populus trichocarpa) produces a variety of volatile and nonvolatile benzenoids involved in various defense responses. However, their biosynthesis in poplar is mainly unresolved. We showed feeding of the poplar leaf beetle (Chrysomela populi) on P. trichocarpa leaves led to increased emission of the benzenoid volatiles benzaldehyde, benzylalcohol, and benzyl benzoate. The accumulation of salicinoids, a group of nonvolatile phenolic defense glycosides composed in part of benzenoid units, was hardly affected by beetle herbivory. In planta labeling experiments revealed that volatile and nonvolatile poplar benzenoids are produced from cinnamic acid (C6–C3). The biosynthesis of C6–C1 aromatic compounds from cinnamic acid has been described in petunia (Petunia hybrida) flowers where the pathway includes a peroxisomal-localized chain shortening sequence, involving cinnamate-CoA ligase (CNL), cinnamoyl-CoA hydratase/dehydrogenase (CHD), and 3-ketoacyl-CoA thiolase (KAT). Sequence and phylogenetic analysis enabled the identification of small CNL, CHD, and KAT gene families in P. trichocarpa. Heterologous expression of the candidate genes in Escherichia coli and characterization of purified proteins in vitro revealed enzymatic activities similar to those described in petunia flowers. RNA interference-mediated knockdown of the CNL subfamily in gray poplar (Populus x canescens) resulted in decreased emission of C6–C1 aromatic volatiles upon herbivory, while constitutively accumulating salicinoids were not affected. This indicates the peroxisomal β-oxidative pathway participates in the formation of volatile benzenoids. The chain shortening steps for salicinoids, however, likely employ an alternative pathway.

Overexpression of bacterial katE gene improves the resistance of modified tomato plant against Fusarium oxysporum f. sp. lycopersici

ABSTRACT Tomato (Solanum lycopersicum L.) yield is severely affected by Fusarium fungal disease. To improve the resistance of tomato against Fusarium oxysporum f. sp. lycopersici (FOL), Escherichia coli katE gene was transformed into two tomato cultivars, namely Castle Rock and Super strain B, via Agrobacterium tumefaciens; the transformation efficiency was 5.6% and 3.5%, respectively. The integration of the katE gene into T0, T1, and T2 transgenic tomato lines was confirmed using PCR. In addition, DNA dot blot technique confirmed the integration of the katE gene into T2 transgenic tomato lines. The RT-PCR analysis confirmed that the katE gene could be expressed normally in the T2 modified lines. Under artificial infection with FOL, the non-modified plants exhibited more severe fungal disease symptoms than those observed in katE overexpression (OE) lines. Our analysis showed that the levels of three defense enzymes, namely superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), were increased during transgenic T2 generation pre-treated with FOL. The bioassay of modified lines revealed that an average of 52.56% of the modified Castle Rock cultivar and 50.28% of the modified Super Strain B cultivar showed resistance under Fusarium infection. These results clearly indicate that the modified tomato plants, in which the katE gene was overexpressed, became more resistant to the infection by FOL than the wild-type plants. Our study has proven that the overexpression of the E. coli katE gene in the OE lines could be utilized to develop and improve the resistance against fungal diseases in the modified crops.

Metagenomic characterization of lysine acetyltransferases in human cancer and their association with clinicopathologic features.

Lysine acetyltransferases (KATs) are a highly diverse group of epigenetic enzymes that play important roles in various cellular processes including transcription, signal transduction, and cellular metabolism. However, our knowledge of the genomic and transcriptomic alterations of KAT genes and their clinical significance in human cancer remains incomplete. We undertook a metagenomic analysis of 37 KATs in more than 10 000 cancer samples across 33 tumor types, focusing on breast cancer. We identified associations among recurrent genetic alteration, gene expression, clinicopathologic features, and patient survival. Loss‐of‐function analysis was carried out to examine which KAT has important roles in growth and viability of breast cancer cells. We identified that a subset of KAT genes, including NAA10, KAT6A, and CREBBP, have high frequencies of genomic amplification or mutation in a spectrum of human cancers. Importantly, we found that 3 KATs, NAA10, ACAT2, and BRD4, were highly expressed in the aggressive basal‐like subtype, and their expression was significantly associated with disease‐free survival. Furthermore, we showed that depletion of NAA10 inhibits basal‐like breast cancer growth in vitro. Our findings provide a strong foundation for further mechanistic research and for developing therapies that target NAA10 or other KATs in human cancer.

Lysine acetylation as drug target in fungi: an underexplored potential in Aspergillus spp.

In recent years, the intensification of the use of immunosuppressive therapies has increased the incidence of invasive infections caused by opportunistic fungi. Considering that, the spread of azole resistance and amphotericin B (AmB) inefficiency against some clinical and environmental isolates has been described. Thus, to avoid a global problem when controlling fungal infections and critical failures in medicine, and food security, new approaches for drug target identification and for the development of new treatments that are more effective against pathogenic fungi are desired. Recent studies indicate that protein acetylation is present in hundreds of proteins of different cellular compartments and is involved in several biological processes, i.e., metabolism, translation, gene expression regulation, and oxidative stress response, from prokaryotes and eukaryotes, including fungi, demonstrating that lysine acetylation plays an important role in essential mechanisms. Lysine acetyltransferases (KATs) and lysine deacetylases (KDACs), the two enzyme families responsible for regulating protein acetylation levels, have been explored as drug targets for the treatment of several human diseases and infections. Aspergilli have on average 8 KAT genes and 11 KDAC genes in their genomes. This review aims to summarize the available knowledge about Aspergillus spp. azole resistance mechanisms and the role of lysine acetylation in the control of biological processes in fungi. We also want to discuss the lysine acetylation as a potential target for fungal infection treatment and drug target discovery.

The Campylobacter jejuni Type VI Secretion System Enhances the Oxidative Stress Response and Host Colonization.

The role of the Type VI secretion system (T6SS) in Campylobacter jejuni is poorly understood despite an increasing prevalence of the T6SS in recent C. jejuni isolates in humans and chickens. The T6SS is a contractile secretion machinery capable of delivering effectors that can play a role in host colonization and niche establishment. During host colonization, C. jejuni is exposed to oxidative stress in the host gastrointestinal tract, and in other bacteria the T6SS has been linked with the oxidative stress response. In this study, comparisons of whole genome sequences of a novel human isolate 488 with previously sequenced strains revealed a single highly conserved T6SS cluster shared between strains isolated from humans and chickens. The presence of a functional T6SS in the 488 wild-type strain is indicated by expression of T6SS genes and secretion of the effector TssD. Increased expression of oxidative stress response genes katA, sodB, and ahpC, and increased oxidative stress resistance in 488 wild-type strain suggest T6SS is associated with oxidative stress response. The role of the T6SS in interactions with host cells is explored using in vitro and in vivo models, and the presence of the T6SS is shown to increase C. jejuni cytotoxicity in the Galleria mellonella infection model. In biologically relevant models, the T6SS enhances C. jejuni interactions with and invasion of chicken primary intestinal cells and enhances the ability of C. jejuni to colonize chickens. This study demonstrates that the C. jejuni T6SS provides defense against oxidative stress and enhances host colonization, and highlights the importance of the T6SS during in vivo survival of T6SS-positive C. jejuni strains.