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.

scholarly journals DNA Array-Based Transcriptional Analysis of Asporogenous, Nonsolventogenic Clostridium acetobutylicum Strains SKO1 and M5

2003 ◽  
Vol 185 (15) ◽  
pp. 4539-4547 ◽  
Author(s):  
Christopher A. Tomas ◽  
Keith V. Alsaker ◽  
Hendrik P. J. Bonarius ◽  
Wouter T. Hendriksen ◽  
He Yang ◽  
...  
Keyword(s):  
Electron Transport ◽  
Clostridium Acetobutylicum ◽  
Large Scale ◽  
Dna Microarrays ◽  
Expression Patterns ◽  
Central Carbon Metabolism ◽  
Transcriptional Analysis ◽  
Dna Arrays ◽  
Major Cluster ◽  
Solvent Formation

ABSTRACT The large-scale transcriptional program of two Clostridium acetobutylicum strains (SKO1 and M5) relative to that of the parent strain (wild type [WT]) was examined by using DNA microarrays. Glass DNA arrays containing a selected set of 1,019 genes (including all 178 pSOL1 genes) covering more than 25% of the whole genome were designed, constructed, and validated for data reliability. Strain SKO1, with an inactivated spo0A gene, displays an asporogenous, filamentous, and largely deficient solventogenic phenotype. SKO1 displays downregulation of all solvent formation genes, sigF, and carbohydrate metabolism genes (similar to genes expressed as part of the stationary-phase response in Bacillus subtilis) but also several electron transport genes. A major cluster of genes upregulated in SKO1 includes abrB, the genes from the major chemotaxis and motility operons, and glycosylation genes. Strain M5 displays an asporogenous and nonsolventogenic phenotype due to loss of the megaplasmid pSOL1, which contains all genes necessary for solvent formation. Therefore, M5 displays downregulation of all pSOL1 genes expressed in the WT. Notable among other genes expressed more highly in WT than in M5 were sigF, several two-component histidine kinases, spo0A, cheA, cheC, many stress response genes, fts family genes, DNA topoisomerase genes, and central-carbon metabolism genes. Genes expressed more highly in M5 include electron transport genes (but different from those downregulated in SKO1) and several motility and chemotaxis genes. Most of these expression patterns were consistent with phenotypic characteristics. Several of these expression patterns are new or different from what is known in B. subtilis and can be used to test a number of functional-genomic hypotheses.

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.

Over-expression of stress protein-encoding genes helps Clostridium acetobutylicum to rapidly adapt to butanol stress

2012 ◽  
Vol 34 (9) ◽  
pp. 1643-1649 ◽  
Author(s):  
Miriam S. Mann ◽  
Zdravko Dragovic ◽  
Georg Schirrmacher ◽  
Tina Lütke-Eversloh
Keyword(s):  
Clostridium Acetobutylicum ◽  
Stress Protein ◽  
Over Expression ◽  
Protein Encoding ◽  
Butanol Stress ◽  
Encoding 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 Genome-wide in silico identification, characterization and transcriptional analysis of the family of growth-regulating factors in common bean (Phaseolus vulgaris L.) subjected to polyethylene glycol-induced drought stress

2017 ◽  
Vol 69 (1) ◽  
pp. 5-14 ◽  
Author(s):  
İlker Büyük ◽  
Sümer Aras
Keyword(s):  
Drought Stress ◽  
Stress Response ◽  
Phaseolus Vulgaris ◽  
Common Bean ◽  
In Silico ◽  
Expression Patterns ◽  
Transcriptional Analysis ◽  
Genome Wide ◽  
The Common ◽  
Root Tissues

According to most recent findings, growth regulating factors (GRFs) are plant-specific transcription factors (TFs) that play important roles in many processes, including abiotic and biotic stress response mechanisms. Completion of the common bean (Phaseolus vulgaris) genome project has provided researchers with the opportunity to identify all GRF genes in this species. With this aim, a genome-wide in silico study was performed and 10 GRF proteins (called PhvGRFs) were identified in the common bean genome. Conserved and mandatory motifs (QLQ and WRC) were confirmed in all identified PhvGRFs and two segmental duplication events were determined. Most of the PhvGRFs were found to be more similar to Arabidopsis thaliana GRFs than to Zea mays GRFs in a phylogenetic tree. According to the expression analysis of 10 PhvGRFs, inversely related expression patterns were observed in the roots of Yakutiye and Zulbiye cultivars based on their capacity to adopt to drought stress. After drought treatment of the Zulbiye cultivar, a drought-sensitive common bean cultivar, PhvGRF1, PhvGRF2, PhvGRF3, PhvGRF5, PhvGRF6, PhvGRF9 and PhvGRF10 genes were upregulated 2- to 4-fold in root tissues, as compared to the untreated control. The trend of PhvGRF1, PhvGRF2, PhvGRF3, PhvGRF5, PhvGRF6, PhvGRF7, PhvGRF9 and PhvGRF10 genes showed a consistent decline of 2- to 6-fold in root tissues of the drought-tolerant Yakutiye cultivar subjected to 24 h of drought stress. We demonstrated that the expression patterns of the identified PhvGRFs correlated with the drought-stress response in a cultivar-specific manner in the common bean. We suggest that members of the GRF family can also be used for genetic engineering applications in the common bean.

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 Intracellular Butyryl Phosphate and Acetyl Phosphate Concentrations in Clostridium acetobutylicum and Their Implications for Solvent Formation

2005 ◽  
Vol 71 (1) ◽  
pp. 530-537 ◽  
Author(s):  
Yinsuo Zhao ◽  
Christopher A. Tomas ◽  
Fredrick B. Rudolph ◽  
Eleftherios T. Papoutsakis ◽  
George N. Bennett
Keyword(s):  
Clostridium Acetobutylicum ◽  
Dry Weight ◽  
Transcriptional Analysis ◽  
Acetyl Phosphate ◽  
Early Exponential Phase ◽  
Stress Genes ◽  
Regulatory Molecule ◽  
Solvent Formation ◽  
Butyrate Kinase ◽  
Second Peak

ABSTRACT It has been suggested (L. H. Harris, R. P. Desai, N. E. Welker, and E. T. Papoutsakis, Biotechnol. Bioeng. 67:1-11, 2000) that butyryl phosphate (BuP) is a regulator of solventogenesis in Clostridium acetobutylicum. Here, we determined BuP and acetyl phosphate (AcP) levels in fermentations of C. acetobutylicum wild type (WT), degenerate strain M5, a butyrate kinase (buk) mutant, and a phosphotransacetylase (pta) mutant. A sensitive method was developed to measure BuP and AcP in the same sample. Compared to the WT, the buk mutant had higher levels of BuP and AcP; the BuP levels were high during the early exponential phase, and there was a peak corresponding to solvent production. Consistent with this, solvent formation was initiated significantly earlier and was much stronger in the buk mutant than in all other strains. For all strains, initiation of butanol formation corresponded to a BuP peak concentration that was more than 60 to 70 pmol/g (dry weight), and higher and sustained levels corresponded to higher butanol formation fluxes. The BuP levels never exceeded 40 to 50 pmol/g (dry weight) in strain M5, which produces no solvents. The BuP profiles were bimodal, and there was a second peak midway through solventogenesis that corresponded to carboxylic acid reutilization. AcP showed a delayed single peak during late solventogenesis corresponding to acetate reutilization. As expected, in the pta mutant the AcP levels were very low, yet this strain exhibited strong butanol production. These data suggest that BuP is a regulatory molecule that may act as a phosphodonor of transcriptional factors. DNA array-based transcriptional analysis of the buk and M5 mutants demonstrated that high BuP levels corresponded to downregulation of flagellar genes and upregulation of solvent formation and stress genes.

scholarly journals Signalling pathways and mechanistic cues highlighted by transcriptomic analysis of primordial, primary, and secondary ovarian follicles in domestic cat

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Shauna Kehoe ◽  
Katarina Jewgenow ◽  
Paul R. Johnston ◽  
Susan Mbedi ◽  
Beate C. Braun
Keyword(s):  
Gene Expression ◽  
Transforming Growth Factor ◽  
Ovarian Follicle ◽  
Mammalian Species ◽  
Expression Patterns ◽  
Ovarian Follicles ◽  
Domestic Cat ◽  
Transcriptional Analysis ◽  
Ovarian Folliculogenesis

AbstractIn vitro growth (IVG) of dormant primordial ovarian follicles aims to produce mature competent oocytes for assisted reproduction. Success is dependent on optimal in vitro conditions complemented with an understanding of oocyte and ovarian follicle development in vivo. Complete IVG has not been achieved in any other mammalian species besides mice. Furthermore, ovarian folliculogenesis remains sparsely understood overall. Here, gene expression patterns were characterised by RNA-sequencing in primordial (PrF), primary (PF), and secondary (SF) ovarian follicles from Felis catus (domestic cat) ovaries. Two major transitions were investigated: PrF-PF and PF-SF. Transcriptional analysis revealed a higher proportion in gene expression changes during the PrF-PF transition. Key influencing factors during this transition included the interaction between the extracellular matrix (ECM) and matrix metalloproteinase (MMPs) along with nuclear components such as, histone HIST1H1T (H1.6). Conserved signalling factors and expression patterns previously described during mammalian ovarian folliculogenesis were observed. Species-specific features during domestic cat ovarian folliculogenesis were also found. The signalling pathway terms “PI3K-Akt”, “transforming growth factor-β receptor”, “ErbB”, and “HIF-1” from the functional annotation analysis were studied. Some results highlighted mechanistic cues potentially involved in PrF development in the domestic cat. Overall, this study provides an insight into regulatory factors and pathways during preantral ovarian folliculogenesis in domestic cat.

scholarly journals Effects of the Bacterial Extract OM-85 on Phagocyte Functions and the Stress Response

1994 ◽  
Vol 3 (2) ◽  
pp. 143-148 ◽  
Author(s):  
S. Baladi ◽  
S. Kantengwa ◽  
Y. R. A. Donati ◽  
B. S. Polla
Keyword(s):  
Stress Response ◽  
Respiratory Burst ◽  
Stress Proteins ◽  
Human Peripheral Blood ◽  
Stress Protein ◽  
Blood Monocytes ◽  
Bacterial Extract ◽  
Heat Shock Stress ◽  
Intracellular Expression ◽  
Glucose Regulated Protein

The effects of the bacterial extract OM-85 on the respiratory burst, intracellular calcium and the stress response have been investigated in human peripheral blood monocytes from normal donors. Activation of the respiratory burst during bacterial phagocytosis has been previously associated with heat shock/stress proteins synthesis. Whereas OM-85 stimulated superoxide production and increased Ca2+mobilization, it fared to induce synthesis of classical HSPs. The lack of stress protein induction was observed even in the presence of iron which potentiates both oxidative injury and stress protein induction during bacterial phagocytosis. However OM-85 induced a 75–78 kDa protein, which is likely to be a glucose regulated protein (GRP78), and enhanced intracellular expression of interleukin-lβ precursor.

scholarly journals Molecular Characterization and Functional Analysis of PR-1-Like Proteins Identified from the Wheat Head Blight Fungus Fusarium graminearum

2018 ◽  
Vol 108 (4) ◽  
pp. 510-520 ◽  
Author(s):  
Shunwen Lu ◽  
Michael C. Edwards
Keyword(s):  
Functional Analysis ◽  
Fusarium Graminearum ◽  
Expression Patterns ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Transcriptional Analysis ◽  
Fungal Genome ◽  
Dimensional Structure ◽  
Head Blight ◽  
Fungal Virulence

The group 1 pathogenesis-related (PR-1) proteins originally identified from plants and their homologs are also found in other eukaryotic kingdoms. Studies on nonplant PR-1-like (PR-1L) proteins have been pursued widely in humans and animals but rarely in filamentous ascomycetes. Here, we report the characterization of four PR-1L proteins identified from the ascomycete fungus Fusarium graminearum, the primary cause of Fusarium head blight of wheat and barley (designated FgPR-1L). Molecular cloning revealed that the four FgPR-1L proteins are all encoded by small open reading frames (612 to 909 bp) that are often interrupted by introns, in contrast to plant PR-1 genes that lack introns. Sequence analysis indicated that all FgPR-1L proteins contain the PR-1-specific three-dimensional structure, and one of them features a C-terminal transmembrane (TM) domain that has not been reported for any stand-alone PR-1 proteins. Transcriptional analysis revealed that the four FgPR-1L genes are expressed in axenic cultures and in planta with different spatial or temporal expression patterns. Phylogenetic analysis indicated that fungal PR-1L proteins fall into three major groups, one of which harbors FgPR-1L-2-related TM-containing proteins from both phytopathogenic and human-pathogenic ascomycetes. Low-temperature sodium dodecyl sulfate polyacrylamide gel electrophoresis and proteolytic assays indicated that the recombinant FgPR-1L-4 protein exists as a monomer and is resistant to subtilisin of the serine protease family. Functional analysis confirmed that deletion of the FgPR-1L-4 gene from the fungal genome results in significantly reduced virulence on susceptible wheat. This study provides the first example that the F. graminearum–wheat interaction involves a pathogen-derived PR-1L protein that affects fungal virulence on the host.

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