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

Abstract

Background Aerobic 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. Methods Antioxidant 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. Results P. 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. Conclusions This 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.

Pseudomonas putida as saviour for troubled Synechococcus elongatus in a synthetic co-culture – Interaction studies based on a multi-OMICs approach

“…However, only one of the two major cellular ROS degrading regulators, SoxR (PP_2060 log2-FC 1.1), was found to be marginally upregulated. Another mechanism in the antioxidant defence is the glutathione metabolism 11,42 , however no regulated genes associated with glutathione could be identified. Nonetheless, on metabolomic level more metabolites belonging to the glutathione metabolism could be found in the co-culture cells (Figure 9E).…”

The long-chain flavodoxin FldX1 improves the biodegradation of 4-hydroxyphenylacetate and 3-hydroxyphenylacetate and counteracts the oxidative stress associated to aromatic catabolism in Paraburkholderia xenovorans

“…Bacteria that inhabit aerobic environments are exposed to the harmful effects of reactive oxygen species (ROS), such as hydrogen peroxide (H 2 O 2 ), superoxide radical (O 2 − ), and hydroxyl radical (OH˙), which are intracellularly generated by the reduction of molecular oxygen [ 1 – 3 ]. ROS may generate cellular damage disrupting iron-cofactor enzymes, and oxidizing DNA, proteins, and lipids [ 3 , 4 ]. Regulation of membrane permeability, antioxidant and repair systems, ROS-scavenging enzymes, and replacement of ROS-sensitive targets by resistant isofunctional enzymes are part of the adaptive mechanisms to tolerate aerobic environments, capable of counteracting the internal oxidation to nontoxic levels [ 3 , 4 ].…”

“…ROS may generate cellular damage disrupting iron-cofactor enzymes, and oxidizing DNA, proteins, and lipids [ 3 , 4 ]. Regulation of membrane permeability, antioxidant and repair systems, ROS-scavenging enzymes, and replacement of ROS-sensitive targets by resistant isofunctional enzymes are part of the adaptive mechanisms to tolerate aerobic environments, capable of counteracting the internal oxidation to nontoxic levels [ 3 , 4 ]. Interestingly, H 2 O 2 is also an anaerobic electron acceptor for E. coli in the absence of molecular oxygen by the respiratory oxidase cytochrome c peroxidase, reducing its toxicity [ 5 ].…”

“…ROS accumulation can occur in the cell when ROS levels exceed the defense mechanisms, entering to a status known as oxidative stress, inducing scavenging enzymes and other general stress proteins [ 2 , 7 ]. These enzymatic systems are induced in bacteria directly by exposure to oxidizing agents, such as H 2 O 2 , superoxide or NO, and indirectly via a general stress response caused by environmental factors and chemical agents [ 4 , 8 – 11 ]. Exposure to environmental stresses can cause a general detriment in cellular growth, performance, survival and fitness, partly related to ROS accumulation and oxidative damage.…”

“…Paraburkholderia xenovorans LB400 is a model bacterium for the degradation of polychlorobiphenyls (PCBs) and diverse aromatic compounds [ 20 – 27 ]. The LB400 genome contains genes encoding for oxygenases, dioxygenases, and hydroxylases, key enzymes involved in the degradation of aromatic compounds, as well as a wide repertoire of genes involved in oxidative stress response [ 4 , 24 ]. Exposure of P. xenovorans LB400 to (chloro)biphenyls ( i.e.…”