Improvement of Thermotolerance of Zymomonas mobilis by Genes for Reactive Oxygen Species-Scavenging Enzymes and Heat Shock Proteins.
Ontology highlight
ABSTRACT:
SUBMITTER: Anggarini S
PROVIDER: S-EPMC7002363 | biostudies-literature | 2019
REPOSITORIES: biostudies-literature
Publications
Improvement of Thermotolerance of <i>Zymomonas mobilis</i> by Genes for Reactive Oxygen Species-Scavenging Enzymes and Heat Shock Proteins.
Anggarini Sakunda S Murata Masayuki M Kido Keisuke K Kosaka Tomoyuki T Sootsuwan Kaewta K Thanonkeo Pornthap P Yamada Mamoru M
Frontiers in microbiology 20200130
Thermotolerant genes, which are essential for survival at a high temperature, have been identified in three mesophilic microbes, including <i>Zymomonas mobilis</i>. Contrary to expectation, they include only a few genes for reactive oxygen species (ROS)-scavenging enzymes and heat shock proteins, which are assumed to play key roles at a critical high temperature (CHT) as an upper limit of survival. We thus examined the effects of increased expression of these genes on the cell growth of <i>Z. mo ...[more]
Similar Datasets
Project description:Zymomonas mobilis ZM4 produces near theoretical yields of ethanol with high specific productivity and recombinant strains are able to ferment both C-5 and C-6 sugars. However, the genetic and physiological basis of the ZM4 response to various industrially-relevant stresses is poorly understood. In this study, the dynamics of ZM4 oxygen stress responses were elucidated by characterizing the transcriptomic and metabolomic profiles of aerobic and anaerobic fermentations using whole-genome microarray analysis and gas chromatography-mass spectrometry. In the absence of oxygen, ZM4 consumed glucose more rapidly, had a higher growth rate, and ethanol was the major end-product. Under aerobic conditions, much less ethanol was observed with other end-products produced, including acetate, lactate, and acetoin. In the early exponential phase, no genes were detected as being significantly differentially expressed between aerobic and anaerobic conditions via microarray analysis. However, microarray analysis of the stationary phase cultures revealed that 166 genes were significantly differentially expressed by more than two-fold. Quantitative-PCR validated the expression values for seventeen genes from different categories of stationary phase microarray data. Transcripts for Entner-Doudoroff pathway genes and gene pdc, encoding a key enzyme leading to the ethanol production, were at least 30-fold more abundant under anaerobic conditions at stationary phase based on quantitative-PCR results. Expression of stress response genes was found to be greater under oxygen stress conditions. GC-MS analysis of stationary phase intracellular metabolites indicated that ZM4 under anaerobic conditions contained lower levels of amino acids, glucose and ED pathway intermediates, whereas metabolites such as ribitol, trehalose, myristic acid, glyceric acid, glucose 6-phosphate, mannose 6-phosphate, and 4-hydroxybutanoic acid were more abundant than under aerobic conditions. Transcriptomic and metabolomic data were consistent with faster glucose consumption and greater ethanol production under anaerobic conditions and suggested gene targets for deletion and improved fermentation. Keywords: Time course/ Stress Response
2008-10-10 | GSE10302 | GEO
Project description:Reactive oxygen species (ROS) are signaling molecules essential for plant responses to abiotic and biotic stimuli as well as for multiple developmental processes. They are produced as byproducts of aerobic metabolism and are affected by adverse environmental conditions. The ROS content is controlled on the side of their production but also by scavenging machinery. Antioxidant enzymes represent a major ROS-scavenging force and are crucial for stress tolerance in plants. Enzymatic antioxidant defense occurs as a series of redox reactions for ROS elimination. Therefore, the deregulation of the antioxidant machinery may lead to the overaccumulation of ROS in plants, with negative consequences both in terms of plant development and resistance to environmental challenges. The transcriptional activation of antioxidant enzymes accompanies the long-term exposure of plants to unfavorable environmental conditions. Fast ROS production requires the immediate mobilization of the antioxidant defense system, which may occur via retrograde signaling, redox-based modifications, and the phosphorylation of ROS detoxifying enzymes. This review aimed to summarize the current knowledge on signaling processes regulating the enzymatic antioxidant capacity of plants.
| S-EPMC7882706 | biostudies-literature
Project description:Zymomonas mobilis ZM4 produces near theoretical yields of ethanol with high specific productivity and recombinant strains are able to ferment both C-5 and C-6 sugars. However, the genetic and physiological basis of the ZM4 response to various industrially-relevant stresses is poorly understood. In this study, the dynamics of ZM4 oxygen stress responses were elucidated by characterizing the transcriptomic and metabolomic profiles of aerobic and anaerobic fermentations using whole-genome microarray analysis and gas chromatography-mass spectrometry. In the absence of oxygen, ZM4 consumed glucose more rapidly, had a higher growth rate, and ethanol was the major end-product. Under aerobic conditions, much less ethanol was observed with other end-products produced, including acetate, lactate, and acetoin. In the early exponential phase, no genes were detected as being significantly differentially expressed between aerobic and anaerobic conditions via microarray analysis. However, microarray analysis of the stationary phase cultures revealed that 166 genes were significantly differentially expressed by more than two-fold. Quantitative-PCR validated the expression values for seventeen genes from different categories of stationary phase microarray data. Transcripts for Entner-Doudoroff pathway genes and gene pdc, encoding a key enzyme leading to the ethanol production, were at least 30-fold more abundant under anaerobic conditions at stationary phase based on quantitative-PCR results. Expression of stress response genes was found to be greater under oxygen stress conditions. GC-MS analysis of stationary phase intracellular metabolites indicated that ZM4 under anaerobic conditions contained lower levels of amino acids, glucose and ED pathway intermediates, whereas metabolites such as ribitol, trehalose, myristic acid, glyceric acid, glucose 6-phosphate, mannose 6-phosphate, and 4-hydroxybutanoic acid were more abundant than under aerobic conditions. Transcriptomic and metabolomic data were consistent with faster glucose consumption and greater ethanol production under anaerobic conditions and suggested gene targets for deletion and improved fermentation. Keywords: Time course/ Stress Response A total of 24 samples were analyzed. These consisted of two times points, (3 hours and 26 hours) under two conditions (anaerobic and aerobic). There were 3 biological replicates and two dye swaps. Each microarray containted one to two probes per predicted coding sequence.
2010-05-18 | E-GEOD-10302 | biostudies-arrayexpress
Project description:BACKGROUND:Global warming threatens rice growth and reduces yields. Proline plays important roles in plant abiotic stress tolerance. Previous research demonstrated that engineering proline metabolism-related genes can enhance tolerance to freezing and salinity in Arabidopsis. OsProDH encodes a putative proline dehydrogenase and is a single copy gene in rice. However, whether OsProDH plays roles in abiotic stress in rice remains unknown. FINDINGS:Quantitative RT-PCR analysis revealed that OsProDH transcript contents were relatively higher in leaf blade and root tissues and the high temperature treatment repressed expression of OsProDH. The predicted OsProDH protein localized in mitochondria. Using the Oryza sativa ssp. japonica cultivar KY131, we generated OsProDH overexpression (OE) lines and knockout mutant lines using the CRISPR/Cas9 (CRI) system. Overexpression of OsProDH decreased proline content, while mutation of OsProDH increased proline content compared with that of KY131. The CRI and OE lines were respectively more resistant and sensitive to heat stress than KY131. Heat stress induced proline accumulation and mutation of OsProDH led to proline overproduction which reduced H2O2 accumulation in the seedlings. CONCLUSIONS:OsProDH negatively regulates thermotolerance in rice. Our study provides a strategy to improve heat tolerance in rice via manipulating proline metabolism.
| S-EPMC7450016 | biostudies-literature
Project description:Development of the strategy known as consolidated bioprocessing (CBP) involves the use of a single microorganism to convert pretreated lignocellulosic biomass to ethanol through the simultaneous production of saccharolytic enzymes and fermentation of the liberated monomeric sugars. In this report, the initial steps toward achieving this goal in the fermentation host Zymomonas mobilis were investigated by expressing heterologous cellulases and subsequently examining the potential to secrete these cellulases extracellularly. Numerous strains of Z. mobilis were found to possess endogenous extracellular activities against carboxymethyl cellulose, suggesting that this microorganism may harbor a favorable environment for the production of additional cellulolytic enzymes. The heterologous expression of two cellulolytic enzymes, E1 and GH12 from Acidothermus cellulolyticus, was examined. Both proteins were successfully expressed as soluble, active enzymes in Z. mobilis although to different levels. While the E1 enzyme was less abundantly expressed, the GH12 enzyme comprised as much as 4.6% of the total cell protein. Additionally, fusing predicted secretion signals native to Z. mobilis to the N termini of E1 and GH12 was found to direct the extracellular secretion of significant levels of active E1 and GH12 enzymes. The subcellular localization of the intracellular pools of cellulases revealed that a significant portion of both the E1 and GH12 secretion constructs resided in the periplasmic space. Our results strongly suggest that Z. mobilis is capable of supporting the expression and secretion of high levels of cellulases relevant to biofuel production, thereby serving as a foundation for developing Z. mobilis into a CBP platform organism.
| S-EPMC2950452 | biostudies-literature
Project description:Acetaldehyde is a valuable product of microbial biosynthesis, which can be used by the chemical industry as the entry point for production of various commodity chemicals. In ethanologenic microorganisms, like yeast or the bacterium Zymomonas mobilis, this compound is the immediate metabolic precursor of ethanol. In aerobic cultures of Z. mobilis, it accumulates as a volatile, inhibitory byproduct, due to the withdrawal of reducing equivalents from the alcohol dehydrogenase reaction by respiration. The active respiratory chain of Z. mobilis with its low energy-coupling efficiency is well-suited for regeneration of NAD+ under conditions when acetaldehyde, but not ethanol, is the desired catabolic product. In the present work, we sought to improve the capacity Z. mobilis to synthesize acetaldehyde, based on predictions of a stoichiometric model of its central metabolism developed herein. According to the model analysis, the main objectives in the course of engineering acetaldehyde producer strains were determined to be: (i) reducing ethanol synthesis via reducing the activity of alcohol dehydrogenase (ADH), and (ii) enhancing the respiratory capacity, either by overexpression of the respiratory NADH dehydrogenase (NDH), or by mutation of other components of respiratory metabolism. Several mutants with elevated respiration rate, decreased alcohol dehydrogenase activity, or a combination of both, were obtained. They were extensively characterized by determining their growth rates, product yields, oxygen consumption rates, ADH, and NDH activities, transcription levels of key catabolic genes, as well as concentrations of central metabolites under aerobic culture conditions. Two mutant strains were selected, with acetaldehyde yield close to 70% of the theoretical maximum value, almost twice the previously published yield for Z. mobilis. These strains can serve as a basis for further development of industrial acetaldehyde producers.
| S-EPMC6868117 | biostudies-literature
Project description:Zymomonas mobilis subsp. mobilis is an efficient ethanol producer with application for industrial production of biofuel. To supplement existing Z. mobilis genomic resources and to facilitate genomic research, we used Oxford Nanopore and Illumina sequencing to assemble the complete genome of the beer spoilage isolate Z. mobilis subsp. mobilis strain NRRL B-1960.
| S-EPMC5532819 | biostudies-literature
Project description:Ascorbate peroxidase (APX) acts indispensably in synthesizing L-ascorbate (AsA) which is pivotal to plant stress tolerance by detoxifying reactive oxygen species (ROS). Enhanced activity of APX has been shown to be a key step for genetic engineering of improving plant tolerance. However it needs a deeper understanding on the maintenance of cellular ROS homeostasis in response to stress. In this study, we identified and characterized an APX (CaAPX) gene from Camellia azalea. Quantitative real-time PCR (qRT-PCR) analysis showed that CaAPX was expressed in all tissues and peaked in immature green fruits; the expression levels were significantly upregulated upon cold and hot stresses. Transgenic plants displayed marked enhancements of tolerance under both cold and heat treatments, and plant growth was correlated with CaAPX expression levels. Furthermore, we monitored the activities of several ROS-scavenging enzymes including Cu/Zn-SOD, CAT, DHAR, and MDHAR, and we showed that stress tolerance was synchronized with elevated activities of ROS-scavenging. Moreover, gene expression analysis of ROS-scavenging enzymes revealed a role of CaAPX to orchestrate ROS signaling in response to temperature stresses. Overall, this study presents a comprehensive characterization of cellular response related to CaAPX expression and provides insights to breed crops with high temperature tolerances.
| S-EPMC5366785 | biostudies-literature
Project description:Zymomonas mobilis is an aerotolerant anaerobe and prolific ethanologen with attractive characteristics for industrial bioproduct generation. However, there is currently insufficient knowledge of the impact that environmental factors have on flux through industrially relevant biosynthetic pathways. Here, we examined the effect of oxygen exposure on metabolism and gene expression in Z. mobilis by combining targeted metabolomics, mRNA sequencing, and shotgun proteomics. We found that exposure to oxygen profoundly influenced metabolism, inducing both transient metabolic bottlenecks and long-term metabolic remodeling. In particular, oxygen induced a severe but temporary metabolic bottleneck in the methyl erythritol 4-phosphate pathway for isoprenoid biosynthesis caused by oxidative damage to the iron-sulfur cofactors of the final two enzymes in the pathway. This bottleneck was resolved with minimal changes in expression of isoprenoid biosynthetic enzymes. Instead, it was associated with pronounced upregulation of enzymes related to iron-sulfur cluster maintenance and biogenesis (i.e., flavodoxin reductase and the suf operon). We also detected major changes in glucose utilization in the presence of oxygen. Specifically, we observed increased gluconate production following exposure to oxygen, accounting for 18% of glucose uptake. Our results suggest that under aerobic conditions, electrons derived from the oxidation of glucose to gluconate are diverted to the electron transport chain, where they can minimize oxidative damage by reducing reactive oxygen species such as H2O2. This model is supported by the simultaneous upregulation of three membrane-bound dehydrogenases, cytochrome c peroxidase, and a cytochrome bd oxidase following exposure to oxygen. IMPORTANCE Microbially generated biofuels and bioproducts have the potential to provide a more environmentally sustainable alternative to fossil-fuel-derived products. In particular, isoprenoids, a diverse class of natural products, are chemically suitable for use as high-grade transport fuels and other commodity molecules. However, metabolic engineering for increased production of isoprenoids and other bioproducts is limited by an incomplete understanding of factors that control flux through biosynthetic pathways. Here, we examined the native regulation of the isoprenoid biosynthetic pathway in the biofuel producer Zymomonas mobilis. We leveraged oxygen exposure as a means to perturb carbon flux, allowing us to observe the formation and resolution of a metabolic bottleneck in the pathway. Our multi-omics analysis of this perturbation enabled us to identify key auxiliary enzymes whose expression correlates with increased production of isoprenoid precursors, which we propose as potential targets for future metabolic engineering.
| S-EPMC6372839 | biostudies-literature
Project description:The application of systems biology tools holds promise for rational industrial microbial strain development. Here, we characterize a Zymomonas mobilis mutant (AcR) demonstrating sodium acetate tolerance that has potential importance in biofuel development. The genome changes associated with AcR are determined using microarray comparative genome sequencing (CGS) and 454-pyrosequencing. Sanger sequencing analysis is employed to validate genomic differences and to investigate CGS and 454-pyrosequencing limitations. Transcriptomics, genetic data and growth studies indicate that over-expression of the sodium-proton antiporter gene nhaA confers the elevated AcR sodium acetate tolerance phenotype. nhaA over-expression mostly confers enhanced sodium (Na(+)) tolerance and not acetate (Ac(-)) tolerance, unless both ions are present in sufficient quantities. NaAc is more inhibitory than potassium and ammonium acetate for Z. mobilis and the combination of elevated Na(+) and Ac(-) ions exerts a synergistic inhibitory effect for strain ZM4. A structural model for the NhaA sodium-proton antiporter is constructed to provide mechanistic insights. We demonstrate that Saccharomyces cerevisiae sodium-proton antiporter genes also contribute to sodium acetate, potassium acetate, and ammonium acetate tolerances. The present combination of classical and systems biology tools is a paradigm for accelerated industrial strain improvement and combines benefits of few a priori assumptions with detailed, rapid, mechanistic studies.
| S-EPMC2890806 | biostudies-literature