scholarly journals Transcriptional Analysis of spo0A Overexpression in Clostridium acetobutylicum and Its Effect on the Cell's Response to Butanol Stress

2004 ◽  
Vol 186 (7) ◽  
pp. 1959-1971 ◽  
Author(s):  
Keith V. Alsaker ◽  
Thomas R. Spitzer ◽  
Eleftherios T. Papoutsakis
Keyword(s):  
Fatty Acid ◽  
Cell Division ◽  
Stress Response ◽  
Stationary Phase ◽  
Clostridium Acetobutylicum ◽  
Differentially Expressed ◽  
Transcriptional Analysis ◽  
Flux Analysis ◽  
Butanol Stress ◽  
Plasmid Control

ABSTRACT Spo0A is the regulator of stationary-phase events and is required for transcription of solvent formation genes in Clostridium acetobutylicum. In order to elucidate the role of spo0A in differentiation, we performed transcriptional analysis of 824(pMSPOA) (a spo0A-overexpressing C. acetobutylicum strain with enhanced sporulation) against a plasmid control strain. DNA microarray data were contrasted to data from a spo0A knockout strain (SKO1) that neither sporulates nor produces solvents. Transcripts of fatty acid metabolism genes, motility and chemotaxis genes, heat shock protein genes, and genes encoding the Fts family of cell division proteins were differentially expressed in the two strains, suggesting that these genes play roles in sporulation and the solvent stress response. 824(pMSPOA) alone showed significant downregulation of many glycolytic genes in stationary phase, which is consistent with metabolic flux analysis data. Surprisingly, spo0A overexpression resulted in only nominal transcriptional changes of regulatory genes (abrB and sigF) whose expression was significantly altered in SKO1. Overexpression of spo0A imparted increased tolerance and prolonged metabolism in response to butanol stress. While most of the differentially expressed genes appear to be part of a general stress response (similar to patterns in two plasmid control strains and a groESL-overexpressing strain), several genes were expressed at higher levels at early time points after butanol challenge only in 824(pMSPOA). Most of these genes were related to butyryl coenzyme A and butyrate formation and/or assimilation, but they also included the cell division gene ftsX, the gyrase subunit-encoding genes gyrB and gyrA, DNA synthesis and repair genes, and fatty acid synthesis genes, all of which might play a role in the immediate butanol stress response, and thus in enhanced butanol tolerance.

scholarly journals Transcriptional Analysis of Butanol Stress and Tolerance in Clostridium acetobutylicum

2004 ◽  
Vol 186 (7) ◽  
pp. 2006-2018 ◽  
Author(s):  
Christopher A. Tomas ◽  
Jeffrey Beamish ◽  
Eleftherios T. Papoutsakis
Keyword(s):  
Stress Response ◽  
Glucose Metabolism ◽  
Clostridium Acetobutylicum ◽  
Expression Patterns ◽  
Stress Protein ◽  
Transcriptional Analysis ◽  
Solvent Tolerance ◽  
Solvent Formation ◽  
Butanol Stress ◽  
Dose Dependent

ABSTRACT The effects of challenges with low (0.25%, vol/vol) and high (0.75%) concentrations of butanol on the growth, glucose metabolism, product formation, and transcriptional program of the solvent-tolerant Clostridium acetobutylicum strain 824(pGROE1) and the plasmid control strain 824(pSOS95del) were used to study solvent tolerance and stress response. Strain 824(pGROE1) was generated by groESL overexpression. The growth of 824(pGROE1) was less inhibited than that of 824(pSOS95del), and 824(pGROE1) was able to metabolize glucose over the entire course of the culture (60 h postchallenge) while glucose metabolism in 824(pSOS95del) lasted 24 h. A comparison of their respective DNA array-based transcriptional profiles identified genes with similar expression patterns (these genes are likely to be part of a general butanol stress response) and genes with opposite expression patterns (these genes are likely to be associated with increased tolerance to butanol). Both strains exhibited a butanol dose-dependent increase in expression of all major stress protein genes, including groES, dnaKJ, hsp18, and hsp90; all major solvent formation genes, including aad, ctfA and -B, adc, and bdhA and -B (an unexpected and counterintuitive finding); the butyrate formation genes (ptb and buk); the butyryl coenzyme A biosynthesis operon genes; fructose bisphosphate aldolase; and a gene with homology to Bacillus subtilis kinA. A dose-dependent decrease in expression was observed for the genes of the major fatty acid synthesis operon (also an unexpected and counterintuitive finding), several glycolytic genes, and a few sporulation genes. Genes with opposite expression kinetics included rlpA, artP, and a gene encoding a hemin permease. Taken together, these data suggest that stress, even when it derives from the solvent product itself, triggers the induction of the solvent formation genes.

Differentially expressed genes after hyper- and hypo-salt stress in the halophilic archaeonMethanohalophilus portucalensis

2010 ◽  
Vol 56 (4) ◽  
pp. 295-307 ◽  
Author(s):  
Chao-Jen Shih ◽  
Mei-Chin Lai
Keyword(s):  
Salt Stress ◽  
Stress Response ◽  
Differentially Expressed Genes ◽  
Transmembrane Protein ◽  
Stress Protein ◽  
Differentially Expressed ◽  
Transcriptional Analysis ◽  
Salt Stress Response ◽  
Salt Shock

Methanohalophilus portucalensis FDF1 can grow over a range of external NaCl concentrations, from 1.2 to 2.9 mol/L. Differential gene expression in response to long-term hyper-salt stress (3.1 mol/L of NaCl) and hypo-salt stress (0.9 mol/L of NaCl) were compared by differential display RT-PCR. Fourteen differentially expressed genes responding to long-term hyper- or hypo-salt stress were detected, cloned, and sequenced. Several of the differentially expressed genes were related to the unique energy-acquiring methanogenesis pathway in this organism, including the transmembrane protein MttP, cobalamin biosynthesis protein, methenyl-H4MPT cyclohydrolase and monomethylamine methyltransferase. One signal transduction histidine kinase was identified from the hyper-salt stress cultures. Moreover, 3 known stress-response gene homologues — the DNA mismatch repair protein, MutS, the universal stress protein, UspA, and a member of the protein-disaggregating multichaperone system, ClpB — were also detected. The transcriptional analysis of these long-term salt stress response and adaptation-related genes for cells immediately after salt stress indicated that the expression of the energy metabolism genes was arrested during hyper-salt shock, while the chaperone clpB gene was stimulated by both hypo- and hyper-salt shock.

scholarly journals Clostridium acetobutylicum bacterial membrane responses to carbon ion beam irradiation perturbations

2020 ◽  
Author(s):  
Yue Gao ◽  
Xiang Zhou ◽  
Miaomiao Zhang ◽  
Yajun Liu ◽  
Xiaopeng Guo ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Membrane Potential ◽  
Cell Membrane ◽  
Cell Surface ◽  
Membrane Permeability ◽  
Clostridium Acetobutylicum ◽  
Surface Hydrophobicity ◽  
Cell Membrane Permeability ◽  
Butanol Tolerance ◽  
Butanol Stress

Abstract Background Clostridium acetobutylicum is an important strain during acetone-butanol-ethanol (ABE) fermentation. However, butanol has toxic effects on cells, limiting the application of ABE fermentation. Accordingly, in this study, we aimed to elucidate the metabolic mechanisms through which Clostridium adapts to butanol stress to facilitate the industrial utilization of Clostridium. Results First, using cell morphology, cell membrane permeability and membrane potential, cell surface hydrophobicity, and cell membrane fatty acid composition analyses in wild-type (ATCC 824) and butanol-tolerant (Y217) strains under butanol stress, we explored the responses in the cell membrane to evaluate the damage caused by butanol poisoning. After 2.0% (v/v) butanol treatment, the extracellular conductivity of ATCC 824 increased, intracellular proteins and nucleotides were released in large quantities, the fluorescein diacetate staining rate decreased, the membrane potential decreased, and the cell membrane permeability increased. Under butanol shock, the cell surface of Y217 cells remained intact, and its butanol tolerance mechanism increased the integrity of cell membrane and reduced leakage of cell contents caused by changed in cell membrane permeability, thereby preventing butanol damage to the cell membrane. When stimulated with butanol, Y217 cells showed reduced surface hydrophobicity, thereby improving cellular tolerance to butanol. A comparison of differences in fatty acid compositions between ATCC 824 and Y217 cell membranes under butanol stress further demonstrated that maintenance of the normal physiological characteristics of cell membranes played important roles in resisting the impact of organic solvents. Conclusions Our findings clarified the changes in physiological and biochemical characteristics of the mutant Y217 cell membrane stimulated with butanol to enhance its tolerance. These results may provide important theoretical guidance for further accelerating the acquisition of bacteria with high butanol tolerance and promoting butanol production. Moreover, our study provided a scientific basis for improving the industrial and environmental adaptability of Clostridium.

scholarly journals Overexpression of groESL in Clostridium acetobutylicum Results in Increased Solvent Production and Tolerance, Prolonged Metabolism, and Changes in the Cell's Transcriptional Program

2003 ◽  
Vol 69 (8) ◽  
pp. 4951-4965 ◽  
Author(s):  
Christopher A. Tomas ◽  
Neil E. Welker ◽  
Eleftherios T. Papoutsakis
Keyword(s):  
Clostridium Acetobutylicum ◽  
Stress Protein ◽  
Transcriptional Analysis ◽  
Primary Metabolism ◽  
Control Strain ◽  
Wild Type ◽  
Solvent Production ◽  
In The Wild ◽  
Altered Cell ◽  
Plasmid Control

ABSTRACT DNA array and Western analyses were used to examine the effects of groESL overexpression and host-plasmid interactions on solvent production in Clostridium acetobutylicum ATCC 824. Strain 824(pGROE1) was created to overexpress the groESL operon genes from a clostridial thiolase promoter. The growth of 824(pGROE1) was inhibited up to 85% less by a butanol challenge than that of the control strain, 824(pSOS95del). Overexpression of groESL resulted in increased final solvent titers 40% and 33% higher than those of the wild type and plasmid control strains, respectively. Active metabolism lasted two and one half times longer in 824(pGROE1) than in the wild type. Transcriptional analysis of 824(pGROE1) revealed increased expression of motility and chemotaxis genes and a decrease in the expression of the other major stress response genes. Decreased expression of the dnaKJ operon upon overexpression of groESL suggests that groESL functions as a modulator of the CIRCE regulon, which is shown here to include the hsp90 gene. Analysis of the plasmid control strain 824(pSOS95del) revealed complex host-plasmid interactions relative to the wild-type strain, resulting in prolonged biphasic growth and metabolism. Decreased expression of four DNA gyrases resulted in differential expression of many key primary metabolism genes. The ftsA and ftsZ genes were expressed at higher levels in 824(pSOS95del), revealing an altered cell division and sporulation pattern. Both transcriptional and Western analyses revealed elevated stress protein expression in the plasmid-carrying strain.

scholarly journals 13C-flux Analysis Reveals NADPH-balancing Transhydrogenation Cycles in Stationary Phase of Nitrogen-starvingBacillus subtilis

2012 ◽  
Vol 287 (33) ◽  
pp. 27959-27970 ◽  
Author(s):  
Martin Rühl ◽  
Dominique Le Coq ◽  
Stéphane Aymerich ◽  
Uwe Sauer
Keyword(s):  
Stationary Phase ◽  
Flux Analysis ◽  
13C Flux Analysis

scholarly journals Phytohormones Regulate the Non-Redundant Response of ω-3 Fatty Acid Desaturases to Low Temperatures in Chorispora Bungeana

2021 ◽  
Author(s):  
Yulan Shi ◽  
Sizhong Yang ◽  
Xiule Yue ◽  
Zhixing Zhao ◽  
Lizhe An
Keyword(s):  
Fatty Acids ◽  
Abscisic Acid ◽  
Fatty Acid ◽  
Stress Response ◽  
Low Temperatures ◽  
Cultured Cells ◽  
Temperature Decrease ◽  
Fatty Acid Desaturases ◽  
Cold Environments ◽  
Chorispora Bungeana

Abstract To explore the contribution of ω-3 fatty acid desaturases (FADs) to cold stress response in a special cryophyte, Chorispora bungeana (C. bungeana), two plastidial ω-3 FAD genes (CbFAD7 and CbFAD8) were cloned and verified in a Arabidopsis fad7fad8 mutant, before being compared with the microsomal ω-3 FAD gene (CbFAD3) on expression profile. Though these genes were expressed in all tested tissues of C. bungeana, CbFAD7 and CbFAD8 have the highest expression in leaves, while CbFAD3 was mostly expressed in non-green tissues. Low temperatures (4, 0 and -4 ℃) resulted in significant increases in trienoic fatty acids (TAs, mainly C18:3), which were consistent with the non-redundant expression of CbFAD3 and CbFAD8 in suspension-cultured cells, and the coordination of CbFAD7 and CbFAD8 in leaves. Furthermore, the contribution of CbFAD8 increased as temperature decrease in the two tissues. Our data revealed that jasmonie acid and brassinosteroids participated in the cold-responsive expression of these genes in both tissues, and the pyhtohormone regulation in leaves was more complicated with the participation of abscisic acid and gibberellin. These results point to the hormone-regulated non-redundant contribution of ω-3 CbFADs to maintain appropriate level of TAs under low temperatures, which help C. bungeana survive in cold environments.

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