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RNA-seq analysis of glucose fermentation by the natural Isopropanol producer Clostridium beijerinckii DSM6423

ABSTRACT: Purpose:C. beijerinckii DSM 6423 is the most cited natural IpOH producer. Improving the natural production of this strain through a targeted approach required a full sequencing and characterization of its genome, together with transcriptomic analyses of its own regulations.The goals of this study are then to evaluate the transcriptional profile (RNA-Seq) of C. beijerinckii DSM6423, a natural isopropanol producer, during a fermentation of glucose in controlled bioreactors. Methods : A RNA-Seq approach was chosen in order to have a timelapse study of DSM 6423 throughout the fermentation process. Three independent duplicate fermentations of DSM 6423 were carried out in bioreactors on three different weeks, showing good reproducibility. On each cultivation, five biomass samples were collected for RNA-Seq analyses.and DNA was eliminated after DNAse I treatment (AM1906, Invitrogen). The 15 resulting RNA samples were sequenced and analyzed using the previously reconstructed genome of DSM 6423. Results: Using a data analysis workflow (TAMARA) developed by the Genoscope platform of Evry, we were able to highlight the transcriptional regulation along the fermentation by calculating the transcription profiles of each gene, using the 3h sample as reference. Clustering was performed using CAST algorithm revealed 8 clusters containing 953 genes and corresponding to genes up-regulated at 6, 8, 11 or 24 hours and gene down-regulated at 6, 8, 11 or 24 hours. Conclusion : Such analyses were carried out in this study and provide useful data to better understand the genetic background and the physiological specificities of C. beijerinckii DSM6423 isopropanol producer. Notably, this work is the first omic study of a natural IBE producer. The data gathered needs time for proper exploitation, but a better understanding of the metabolic pathways and various genes involved opens the door for future targeted approaches.

ORGANISM(S): Clostridium beijerinckii

PROVIDER: GSE100024 | GEO | 2018/03/05

REPOSITORIES: GEO

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Project description:Furfural is the prevalent microbial inhibitor generated during pretreatment and hydrolysis of lignocellulosic biomass to monomeric sugars, but the molecular response of Clostridium beijerinckii NCIMB 8052 to this compound is unknown. To discern the effect of furfural on C. beijerinckii and to gain insights into the molecular mechanisms of action and detoxification, we studied the physiological changes of furfural-stressed cultures during acetone-butanol-ethanol (ABE) fermentation, and profiled differentially expressed genes by genome-wide transcriptional analysis. C. beijerinckii exposed to furfural stress during the acidogenic growth phase produced 13% more ABE than the unstressed control. The growth and ABE by C. beijerinckii ceased following exposure to furfural stress during the solventogenic growth phase. By comparing gene expression of furfural-stressed cultures to that of the unstressed control, at both the acidogenic and solventogenic phases, we ascertained that furfural induces expression of several genes including those that code for heat shock proteins, redox enzymes and cofactor associated proteins, and ATP-binding cassette transporters, and represses genes belonging to the phosphotransferase system, two-component system, chemotaxis and cell motility. Based on these results, we discuss the underpinning for furfural-mediated change in ABE fermentation by the solventogenic Clostridium species. C. beijerinckii 8052 pre-culture was incubated anaerobically to attain acidogenic or solventogenic growth phase. The culture was then subdivided into two bottles. One bottle was challenged with furfural and the other bottle was left unchallenged. After 3 h growth, C. beijerinckii 8052 samples were collected, during which the original concentration of furfural in the growth medium was reduced to more than half. Total RNA was isolated, purified, converted to enriched mRNA, and dye-coupled (Alexa Fluor 555) complementary cRNA. This was followed by hybridization and microarray data analysis.

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RNA-seq analysis of glucose fermentation by the natural Isopropanol producer Clostridium beijerinckii DSM6423

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