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The transcriptional pattern of Clostridium acetobutylicum following the downregulation of CAC0654 using antisense RNA

ABSTRACT: Little is known regarding the entry into and the regulation of genes during the sporulation process of clostridia. Using C. acetobutylicum as a model organism we designed an antisense RNA construct targeting the CAC0654 gene to investigate the role of this gene during sporulation and solvent formation. Samples were taken throughout the growth phase and compared to a plasmid control strain to elucidate the transcriptional changes induced by the downregulation of the two-component system CAC0653 and CAC0654

ORGANISM(S): Clostridium acetobutylicum Clostridium acetobutylicum ATCC 824

PROVIDER: GSE14590 | GEO | 2009/01/29

SECONDARY ACCESSION(S): PRJNA111711

REPOSITORIES: GEO

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The transcriptional pattern of Clostridium acetobutylicum following the downregulation of CAC0654 using antisense RNA

Project description:Little is known regarding the entry into and the regulation of genes during the sporulation process of clostridia. Using C. acetobutylicum as a model organism we designed an antisense RNA construct targeting the CAC0654 gene to investigate the role of this gene during sporulation and solvent formation. Samples were taken throughout the growth phase and compared to a plasmid control strain to elucidate the transcriptional changes induced by the downregulation of the two-component system CAC0653 and CAC0654 Samples were taken from a Bioflow II reactor at approximately 9, 12, 13.5, 15, 17, and 19 hrs after bioreactor inoculation containing strain 824(p654as). Two slides per timepoint were hybridized on a dye swap configuration. The control mRNA was generated by combining samples taken at 9, 12, 13.5, 15, 17, and 19 hrs hrs after bioreactor inoculation of the control plasmid strain, 824(pSOS95del). The replicate slide from the 12 hour sample showed high deviations and was discarded from the series.

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CcpA, a Pleiotropic Key Regulator in Butanol-producing Clostridium acetobutylicum ATCC 824

2012-08-30 | GSE33364 | GEO

Global transcriptional changes of Clostridium acetobutylicum cultures with increased butanol-acetone ratios

Project description:Artificial electron carriers have been widely used to shift the solvent ratio towards butanol in acetone-butanol-ethanol (ABE) fermentation of solventogenic clostridia according to decreased hydrogen production. In this study, first insights on the molecular level were gained to explore the effect of methyl viologen addition to cultures of Clostridium acetobutylicum. Employing batch fermentation in mineral salts medium, the butanol:acetone ratio was successively increased from 2.3 to 12.4 on a 100 ml scale in serum bottles and from 1.4 to 16.5 on a 1,300 ml scale in bioreactors, respectively. The latter cultures were used for DNA microarray analyses to provide new information on the transcriptional changes referring to methyl viologen exposure and thus, exhibing gene expression patterns according to the manipulation of the cellular redox balance.

2012-03-31 | E-MEXP-3417 | biostudies-arrayexpress

Regulon prediction of the sporulation factors Spo0A, SigF, SigE, and SigG in Clostridium acetobutylicum ATCC 824

Project description:Clostridium acetobutylicum, the endospore-forming anaerobe best known for its ABE (acetone-butanol-ethanol) fermentation, has received renewed attention recently for the biological production of butanol, both for bulk chemical production and as a potential biofuel. With butanol production in mind, most of the recent research on C. acetobutylicum has focused on increasing butanol production, tolerance to butanol, and optimizing it for various substrates. However, an equally important trait, though less understood, is its sporulation program, which it coupled to solvent formation. The model organism for endospore formation is Bacillus subtilis, but significant physiological, metabolic, and genomic differences exist between the two organisms. Despite these differences, the major sporulation-related transcription/sigma factors are conserved between the two species. These transcription/sigma factors are activated in a cascade manner such that Spo0A becomes active first, followed by σF, then σE, then σG, and finally σK. The goal of this study is to determine the regulons of 4 of these transcription/sigma factors (Spo0A, σF, σE, and σG) and compare them to those in B. subtilis. To accomplish this goal, individual mutant strains were created for Spo0A, σF, σE, and σG, in which the transcription/sigma factor is silenced. These mutants were then compared transcriptionally using microarrays to determine the regulon of each transcription/sigma factor. To help avoid false positives, comparisons were made between strains in which the downstream transcription/sigma factor is silenced (e.g., for the Spo0A regulon, the Spo0A mutant was compared against the σF mutant and the σE mutant, since they are both upregulated by Spo0A) rather than just the WT. For each regulon, 4 timepoints were taken, since it is very difficult to synchronize sporulation in C. acetobutylicum cultures, and dye-swaps were prepared for each timepoint.

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Identification of a sensor histidine kinase (BfcK) controlling biofilm formation in Clostridium acetobutylicum

Project description:Clostridium acetobutylicum has been extensively exploited to produce biofuels and solvents and its biofilm could dramatically improve its productivities. However, genetic control of C. acetobutylicum biofilm has not been dissected so far. Here, a total of 24 disruptants were finally obtained over several years of attempts. Biofilm formation and physiological phenotypes were characterized for these disruptants and most of them showed robust biofilm formation still, or showed both impaired biofilm formation and cell growth. Only a mutant with a disputed histidine kinase gene (CA_C2730, designated bfcK in this study) abolished biofilm formation without impaired cell growth or solvent production. Phosphoproteomic analysis revealed that bfcK could control C. acetobutylicum flagellar motility at both translational and post-translational (protein phosphorylation) levels. The bfcK also showed apparent regulation of a serine/threonine protein kinase (encoded by CA_C0404) which was involved in protein secretion. Based on these findings, possible bfcK-based mechanisms for biofilm formation in C. acetobutylicum were proposed.

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CcpA, a Pleiotropic Key Regulator in Butanol-producing Clostridium acetobutylicum ATCC 824

Project description:Clostridium acetobutylicum is a typical bacterium of major importance to industrial butanol production. In order to dissect the regulatory network pertaining to the industrial application of this bacterium, catabolite control protein A (CcpA) was investigated for its global function by DNA microarray.It showed that CcpA of C. acetobutylicum controls hundreds of genes, not only carbon metabolism, but also solvent production and sporulation in the life cycle.The results here demonstrated that CcpA is an important pleiotropic regulator related to some specific physiological and biochemical process in butanol-producing C. acetobutylicum. In order to enable a global understanding of the regulatory roles of CcpA when fermenting mixed sugars, which is of great significance in utilization of lignocellulosic hydrolysates, D-glucose plus D-xylose were used as the carbon sources in fermentation for microarray analysis. Microarray analysis was performed at four time points:the time point M and L were chosen both in acidogenic phase, while the time point T and S were chosen in shift phase (from acidogenesis to solventogenesis) and solventogenic phase, respectively.One-color microarray assays were performed.Raw data were normalized by Quantile algorithm, Gene Spring Software 11.0 (Agilent technologies, Santa Clara, CA, US). The ratio of transcript level between wildtype and mutant can been achieved using the formula: 2^(value of wildtype)/2^(value of ccpA mutant).

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