Comparative transcriptomics of Clostridium thermocellum wild type (Δhpt) and Type I glutamine synthetase (Clo1313_2031) deletion strains using RNA-seq
Ontology highlight
ABSTRACT:
ORGANISM(S): Acetivibrio thermocellus DSM 1313
PROVIDER: GSE78219 | GEO | 2016/05/31
SECONDARY ACCESSION(S): PRJNA312946
REPOSITORIES: GEO
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Project description:RNA-seq analysis revealed that genes involved in urea uptake and metabolism were significantly upregulated in the Clo1313_2031 deletion strain, suggesting that deletion of Clo1313_2031 mimics nitrogen starvation in C. thermocellum. Additionally, genes encoding the RNF-complex were also more highly expressed and in turn may have a potential role in increasing ethanol production. Samples for RNA-seq were taken from mid-exponential phase (OD ~ 0.33) batch cultures grown in MTC on 4.5 g/l cellobiose
2016-05-31 | E-GEOD-78219 | biostudies-arrayexpress
Project description:Clostridium thermocellum is a Gram-positive, anaerobic, thermophilic bacterium that ferments cellulose into ethanol. It is a candidate industrial consolidated bioprocess (CBP) biocatalyst for lignocellulosic bioethanol production to produce bioethanol directly from cellulosic biomass. However, few transcriptomic studies have been reported so far for C. thermocellum using biomass as carbon source. In this study, samples were taken from exponential and stationary phases of C. thermocellum cells growing in MTC media with pretreated switchgrass as carbon source, and transcriptomic profiling change of C. thermocellum during different growth phase was investigated using both expression array and tiling array. This study will help the understanding of gene expression of C. thermocellum using cellulosic biomass as carbon source and the knowledge will facilitate future metabolic engineering effort for strain improvement.
2014-05-08 | GSE26926 | GEO
Project description:Clostridium thermocellum is a candidate for cellulosic ethanol production. It expresses enzymes for both cellulose solubilization and its fermentation to produce lignocellulosic ethanol. Understanding how this organism regulates gene expression is of importance for developing a better fundamental understanding of this industrial relevant bacterium. We are primarily interested in gene regulation by three predicted LacI regulators. A previous report of one LacI regulator in C. thermocellum ATCC27405 (Cthe_2808) indicated this was a transcriptional repressor of neighboring genes with repressor activity relieved in the presence of laminaribiose (a β-1,3 disaccharide). The current work is aimed at understanding if a homolog of this LacI transcriptional regulator in C. thermocellum DSM1313 and two others putatively characterized as LacI regulators are in fact global regulatory proteins with extensive regulons or, if like the majority of LacI transcription factors, the regulation is limited to local genomic regions. Each of the three LacI regulators was deleted in C. thermocellum DSM1313. Genome re-sequencing was used to confirm the deletion and ensure nonspecific recombination events were avoided during the gene deletion strategy. Growth of each strain on cellobiose was unaffected by any of the gene deletions under pH controlled fermentations. Global gene expression patterns taken at mid-log phase for each strain identified glycoside hydrolase genes encoding hemicellulases, including cellulosomal enzymes, that were highly up-regulated (up to 9 fold) in the absence of each LacI regulator. Thus suggesting these were repressed under wild type conditions. Electrophoretic mobility shift assays have confirmed LacI transcription factor binding to specific regions of gene promoters with putative motifs ranging from 16-18 bp. Work is ongoing to confirm the specific binding motif recognized by the two previously un-characterized transcription factors and assess the occurrence of this motif in the C. thermocellum DSM1313 genome. The identification of LacI repressor activity on hemicellulose gene expression is a key result of this work and will add to the small body of existing literature on the area of gene regulation in C. thermocellum.
2015-10-31 | GSE68423 | GEO
Project description:Clostridium thermocellum is a Gram-positive, anaerobic, thermophilic bacterium that ferments cellulose into ethanol. It is a candidate industrial consolidated bioprocess (CBP) biocatalyst for lignocellulosic bioethanol production. However, C. thermocellum is relatively sensitive to ethanol compared to yeast. Previous studies have investigated the membrane and protein composition of wild-type and ethanol tolerant strains, but relatively little is known about the genome changes associated with the ethanol tolerant C. thermocellum strain. In this study, C. thermocellum cultures were grown to mid-exponential phase and then either shocked with the supplementation of ethanol to a final concentration of 3.95 g/L (equal to 0.5% [v/v]) or were untreated. Samples were taken pre-shock and 2, 12, 30, 60, 120, 240 min post-shock for multiple systems biology analyses. The addition of ethanol dramatically reduced the C. thermocellum growth and the final cell density was approximately half of the control fermentations, with concomitant reductions in substrate consumption in the treated cultures. The response of C. thermocellum to ethanol was dynamic and involved more than six hundred genes that were significantly and differentially expressed between the different conditions over time and every functional category was represented. Cellobiose was accumulated within the ethanol-shocked C. thermocellum cells, as well as the sugar phosphates such as fructose-6-P and cellobiose-6-P. The comparison and correlation among intracellular metabolites, proteomic and transcriptomics profiles as well as the ethanol effects on cellulosome, hydrogenase glycolysis and nitrogen metabolism are discussed, which led us to propose that C. thermocellum may utilize the nitrogen metabolism to bypass the arrested carbon metabolism in responding to ethanol stress shock, and the nitrogen metabolic pathway and redox balance may be the key target for improving ethanol tolerance and production in C. thermocellum.
2012-08-16 | GSE25236 | GEO
Project description:Clostridium thermocellum is a candidate for cellulosic ethanol production. It expresses enzymes for both cellulose solubilization and its fermentation to produce lignocellulosic ethanol. Understanding how this organism regulates gene expression is of importance for developing a better fundamental understanding of this industrial relevant bacterium. We are primarily interested in gene regulation by three predicted LacI regulators. A previous report of one LacI regulator in C. thermocellum ATCC27405 (Cthe_2808) indicated this was a transcriptional repressor of neighboring genes with repressor activity relieved in the presence of laminaribiose (a β-1,3 disaccharide). The current work is aimed at understanding if a homolog of this LacI transcriptional regulator in C. thermocellum DSM1313 and two others putatively characterized as LacI regulators are in fact global regulatory proteins with extensive regulons or, if like the majority of LacI transcription factors, the regulation is limited to local genomic regions. Each of the three LacI regulators was deleted in C. thermocellum DSM1313. Genome re-sequencing was used to confirm the deletion and ensure nonspecific recombination events were avoided during the gene deletion strategy. Growth of each strain on cellobiose was unaffected by any of the gene deletions under pH controlled fermentations. Global gene expression patterns taken at mid-log phase for each strain identified glycoside hydrolase genes encoding hemicellulases, including cellulosomal enzymes, that were highly up-regulated (up to 9 fold) in the absence of each LacI regulator. Thus suggesting these were repressed under wild type conditions. Electrophoretic mobility shift assays have confirmed LacI transcription factor binding to specific regions of gene promoters with putative motifs ranging from 16-18 bp. Work is ongoing to confirm the specific binding motif recognized by the two previously un-characterized transcription factors and assess the occurrence of this motif in the C. thermocellum DSM1313 genome. The identification of LacI repressor activity on hemicellulose gene expression is a key result of this work and will add to the small body of existing literature on the area of gene regulation in C. thermocellum. Four strains of C. thermocellum DSM1313 including a parental strain (Îhpt) and three others with deletion in lacI transcription factors were characterised by RNA-seq. The strains were grown in 1L bioreactors, cells (from 50 ml samples) were harvested at mid exponential, late exponential and 30 min after acid production halted during the transition to stationary phase. The growth studies were performed in triplicate and thirty six samples were analyzed by RNA-sequencing.
2015-10-31 | E-GEOD-68423 | biostudies-arrayexpress
Project description:Differential RNA-Seq analyses to investigate the basis for metabolic inhibition of Clostridium thermocellum M1570 by xylose. The M1570 strain was developed in the C. thermocellum DSM 1313 Δhpt background strain. Lactate dehydrogenase (Ldh) and phosphotransacetylase (Pta) genes are deleted (Argyros DA, Tripathi SA, Barrett TF, Rogers SR, Feinberg LF, Olson DG, Foden JM, Miller BB, Lynd LR, Hogsett DA, Caiazza NC: High ethanol titers from cellulose by using metabolically engineered thermophilic, anaerobic microbes. Appl Environ Microbiol 2010, 77:8288-8294 )
2016-10-31 | GSE74918 | GEO
Project description:Clostridium thermocellum is a promising CBP candidate organism capable of directly converting lignocellulosic biomass to ethanol. Low yields, productivities and growth inhibition prevent industrial deployment of this organism for commodity fuel production. Symptoms of potential redox imbalance such as incomplete substrate utilization, and fermentation products characteristic of overflow metabolism, have been observed during growth. This perceived redox imbalance may be in part responsible for the mentioned bioproductivity limitations. Toward better understanding the redox metabolism of C. thermocellum, we analyzed gene expression, using microarrays, during addition of two stress chemicals (methyl viologen and hydrogen peroxide) which we observed to change fermentation redox potential.
2015-07-29 | GSE71465 | GEO
Project description:Clostridium thermocellum is a Gram-positive, anaerobic, thermophilic bacterium that ferments cellulose into ethanol. It is a candidate industrial consolidated bioprocess (CBP) biocatalyst for lignocellulosic bioethanol production to produce bioethanol directly from cellulosic biomass. However, few transcriptomic studies have been reported so far for C. thermocellum using biomass as carbon source. In this study, samples were taken from exponential and stationary phases of C. thermocellum cells growing in MTC media with pretreated switchgrass as carbon source, and transcriptomic profiling change of C. thermocellum during different growth phase was investigated using both expression array and tiling array. This study will help the understanding of gene expression of C. thermocellum using cellulosic biomass as carbon source and the knowledge will facilitate future metabolic engineering effort for strain improvement. [HX12 expression array]: A eleven array study using total RNA recovered from wild-type cultures of Clostridium thermocellum at different growth phase of T2 and T3 with switchgrass as carbon source. Two biological replicates used for each phase. [3Plex tiling array]: A six array study using total RNA recovered from wild-type cultures of Clostridium thermocellum at different growth phase of T2 and T3 with switchgrass as carbon source. Two biological replicates used for each phase.
2014-05-08 | E-GEOD-26926 | biostudies-arrayexpress
Project description:The thermophilic anaerobe Clostridium thermocellum is a candidate consolidated bioprocessing (CBP) biocatalyst for cellulosic ethanol production. It expresses enzymes for both cellulose solubilization and its fermentation to produce lignocellulosic ethanol. To gain insights into the C. thermocellum genes, using an updated version of the C. thermocellum ATCC 27405 genome annotation, that are required for specific growth on the cellulosic feedstocks of either pretreated switchgrass or Populus, duplicate fermentations were conducted with a 5 g/L solid substrate loading. High quality RNA was extracted using a method we report for C. thermocellum grown on solid substrates. Transcriptome profiles were obtained at two time points during actively growing fermentations (12 h and 37 h post inoculation). A comparison of two transcriptomic analytical techniques, microarray and RNAseq, was performed and the data analyzed for statistical significance. When thresholds for genes passing significance of FDR>0.05 were applied, microarray (2351 genes) had a greater number of significant genes relative to RNA-seq (280 genes when normalized by KDMM). When a 2-fold difference in expression threshold was applied, seventy-three genes were significantly differentially expressed in common between the two techniques. We identified genes differentially expressed when C. thermocellum ATC 27405 was grown on the two biomass substrates, with two putative efflux/transport systems highly differentially regulated (>5-fold). This study has revealed consistency between these two transcriptomics analytical platforms that gives confidence in our switch from the DNA microarray platform to an RNAseq based platform for routine transcriptomics analyses.
2013-05-16 | GSE47010 | GEO
Project description:Clostridium thermocellum is a Gram-positive, anaerobic, thermophilic bacterium that ferments cellulose into ethanol. It is a candidate industrial consolidated bioprocess (CBP) biocatalyst for lignocellulosic bioethanol production. However, C. thermocellum is relatively sensitive to ethanol compared to yeast. Previous studies have investigated the membrane and protein composition of wild-type and ethanol tolerant strains, but relatively little is known about the genome changes associated with the ethanol tolerant C. thermocellum strain. In this study, C. thermocellum cultures were grown to mid-exponential phase and then either shocked with the supplementation of ethanol to a final concentration of 3.95 g/L (equal to 0.5% [v/v]) or were untreated. Samples were taken pre-shock and 2, 12, 30, 60, 120, 240 min post-shock for multiple systems biology analyses. The addition of ethanol dramatically reduced the C. thermocellum growth and the final cell density was approximately half of the control fermentations, with concomitant reductions in substrate consumption in the treated cultures. The response of C. thermocellum to ethanol was dynamic and involved more than six hundred genes that were significantly and differentially expressed between the different conditions over time and every functional category was represented. Cellobiose was accumulated within the ethanol-shocked C. thermocellum cells, as well as the sugar phosphates such as fructose-6-P and cellobiose-6-P. The comparison and correlation among intracellular metabolites, proteomic and transcriptomics profiles as well as the ethanol effects on cellulosome, hydrogenase glycolysis and nitrogen metabolism are discussed, which led us to propose that C. thermocellum may utilize the nitrogen metabolism to bypass the arrested carbon metabolism in responding to ethanol stress shock, and the nitrogen metabolic pathway and redox balance may be the key target for improving ethanol tolerance and production in C. thermocellum. A thirty array study using total RNA recovered from wild-type cultures of Clostridium thermocellum at different time points of 0, 12, 30, 60, 120, and 240 min post-inoculation with 3.95 g/L [0.5% (v/v)] treatment compred to that of control without ethanol supplementation. Two biological replicates for treatment and control condition.
2012-08-16 | E-GEOD-25236 | biostudies-arrayexpress