Project description:Caldicellulosiruptor saccharolyticus is an extremely thermophilic, gram-positive anaerobe which ferments a broad range of substrates to mainly acetate, CO2, and hydrogen gas (H2). Its high hydrogen-producing capacity make this bacterium an attractive candidate for microbial biohydrogen production. However, increased H2 levels tend to inhibit hydrogen formation and leads to the formation of other reduced end products like lactate and ethanol. To investigate the organismM-bM-^@M-^Ys strategy for dealing with elevated H2 levels and to identify alternative pathways involved in the disposal of the reducing equivalents, the effect of the hydrogen partial pressure (PH2) on fermentation performance was studied. For this purpose cultures were grown under high and low PH2 in a glucose limited chemostat setup. Transcriptome analysis revealed the up-regulation of genes involved in the disposal of reducing equivalents under high PH2, like lactate dehydrogenase and alcohol dehydrogenase as well as the NADH-dependent and ferredoxin-dependent hydrogenases. These findings were in line with the observed shift in fermentation profiles from acetate production under low PH2 to a mixed production of acetate, lactate and ethanol under high PH2. In addition, differential transcription was observed for genes involved in carbon metabolism, fatty acid biosynthesis and several transport systems. The presented transcription data provides experimental evidence for the involvement of the redox sensing Rex protein in gene regulation under high PH2 cultivation conditions. Overall, these findings indicate that the PH2 dependent changes in the fermentation pattern of C. saccharolyticus are, in addition to the known regulation at the enzyme/metabolite level, also regulated at the transcription level. Two conditions: low H2 partial pressure and high H2 partial pressure, both at steady state growth were harvested for a dye-flip microarray experimental design. Biological replicates were harvested for both conditions and combined prior to cDNA synthesis. Both conditions were labeled with cy3 and cy5 dyes allowing for a technical replicate of hybridization in addition to the biological replicates.

Project description:【Objective】 The objective of this study is to study the transcriptome regulation mechanism of F. graminearum under different pH stress conditions, analyze the gene expression level and its differences, and explore the metabolic pathways related to the anti-stress response of F. graminearum cells under acidic or alkaline conditions, and reveal how F. graminearum actively regulates intracellular metabolism and synthesis processes to adapt to the changes of extracellular pH environment. 【Method】 F. graminearum was cultured in PDB (potato dextrose broth) medium with initial pH of 4.5, 6.5 and 8.0 for 48 h, and the total RNA of the strain was extracted to construct a cDNA library. Transcriptome sequencing and bioinformatics techniques were used to identify the related differentially expressed genes (DEGs), and the metabolic pathways involved were further analyzed. 【Result】 A total of 4283 DEGs were detected under acidic conditions, of which 2232 were up-regulated and 2252 were down-regulated. Under alkaline conditions, there were a total of 498 DEGs, of which 269 were up-regulated and 229 were down-regulated. The results of Gene Ontology (GO) functional enrichment analysis showed that 211 GO terms were significantly enriched and 72 were down-regulated under acidic conditions. There were 33 GO terms that were revised upwards and 40 downwards under alkaline conditions. The results of KEGG encyclopedia of genes and Genomes (Kyoto Encyclopedia of Genes and Genomes) enrichment analysis showed that 22 pathways were significantly enriched and 32 pathways were down-regulated under acidic conditions. There were 8 up-regulated pathways and 13 down-regulated pathways under alkaline conditions. The expression of membrane transporters and hydrolysis of carbohydrate compounds and other related genes were up-regulated, and the expression of genes related to protein metabolism was down-regulated, which assisted F. graminearum cells to adapt to changes in the external environment. At the same time, F. graminearum maintained the internal environment balance of its own cells by reducing secondary metabolism and amino acid metabolism under acidic and alkaline conditions, respectively, so as to resist extracellular pH stress. 【Conclusion】 In the acidic environment, Fusarium graminearum adapts to the changes in the extracellular environment by promoting the production of riboprotein complexes and secondary metabolism. In an alkaline environment, Fusarium graminearum cells respond to and sense external stresses through amino acid metabolism. The analysis of the metabolic pathways of F. graminearum cells provides important gene expression data for the response of F. graminearum to different pH environments, and the results of this study are helpful to understand the pathogenesis of F. graminearum.

Project description:Thermoacidophilic archaea are found in heavy metal-rich environments and, in some cases, these microorganisms are causative agents of metal mobilization through cellular processes related to their bioenergetics. Given the nature of their habitats, these microorganisms must deal with the potentially toxic effect of heavy metals. Here, we show that two thermoacidophilic Metallosphaera species with nearly identical (99.99%) genomes differed significantly in their sensitivity and reactivity to uranium. M. prunae, isolated from a smoldering heap on a uranium mine in Thuringen, Germany, could be viewed as a M-bM-^@M-^\spontaneous mutantM-bM-^@M-^] of M. sedula, an isolate from Pisciarelli solfatara near Naples, Italy. M. prunae tolerated U3O8 and U(VI) to a much greater extent than M. sedula. Within 15 minutes following exposure to M-bM-^@M-^\U(VI) shockM-bM-^@M-^], M. sedula, and not M. prunae, exhibited transcriptomic features associated with severe stress response. Furthermore, within 15 minutes post-U(VI) shock, M. prunae, and not M. sedula, showed evidence of substantial degradation of cellular RNA. This suggested that transcriptional and translational processes were aborted as a dynamic mechanism for resisting U toxicity; by 60 minutes post-U(VI) shock, RNA integrity in M. prunae recovered, and known modes for heavy metal resistance were activated. In addition, M. sedula rapidly oxidized solid U3O8 to soluble U(VI) for bioenergetic purposes, a chemolithoautotrophic feature not previously reported. M. prunae, however, did not solubilize solid U3O8 to any significant extent, thereby not exacerbating U(VI) toxicity. These results point to uranium extremophily as an adaptive, rather than intrinsic, feature for Metallosphaera species, driven by environmental factors. The study comprises 9 Samples, described in detail below. MprAU_MseAU: Transcriptional analysis of the response of Metallosphaera prunae (Mpr) and Metallosphaera sedula(Mse) to chemolithoautotrophic conditions (0.1 wt% Uranium octaoxide with CO2 supplementation in headspace). This experiment was done to identify the key terminal oxidases which responded to a Uranium oxide while doing inter-species comparison between Mpr and Mse. Transcriptional response of the terminal oxidase clusters proved that certain key genes play a role in the vastly different physiologies of these two species. MprN_MprU60: Transcriptional analysis of the response of Metallosphaera prunae (Mpr) to 60 min of Uranium shock. This experiment was done to analyze the differential transcription of Mpr cells challenged with 1 mM uranyl acetate shock (U shock) compared to normal growth. The Uranium cultures were harvested 60 min after the shock. MprN_MseN: Differential transcription of Metallosphaera species under normal growth conditions. This experiment was done to analyze the differential transcription of Mpr cells compared with Mse cells at mid log phase. MprN_MprU3h: Transcriptional response of Metallosphaera prunae (Mpr) to 3h of Uranium shock compared to normal growth. This experiment was done to analyze the differential transcription of Mpr cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 3 h after the shock. MseN_MseU15: Transcriptional response of Metallosphaera sedula (Mse) to 15 min of Uranium shock. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) compared to normal growth. The Uranium cultures were harvested 15 min after the shock. MseN_MseU60: Transcriptional response of Metallosphaera sedula to 60 min of Uranium shock. Mse cells were grown upto mid log phase after which the cells were subjected to U shock and harvested 60 min later. Biological repeats were done for both experimental conditions. MseN_MseU3h: Transcriptional response of Metallosphaera sedula (Mse) to 3h of Uranium shock compared to normal growth. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 3 h after the shock. MseU15_MseU60: Transcriptional response of Metallosphaera sedula to 15 min of Uranium shock compared with 60 min of Uranium shock. This experiment was done to analyze the differential transcription of Mse cells challenged with 1 mM uranyl acetate shock (U shock) . The Uranium cultures were harvested 15 min and 60 min after the shock. MprU3h_MseU3h: Differential transcription of Metallosphaera cells under Uranium shock. This experiment was done to analyze the differential transcription of Metallosphaera sedula (Mse) and Metallosphaera prunae (Mpr) challenged with 1 mM uranyl acetate.

			Action	DRS
	SRR1163503.fastq.gz	Fastqsanger.gz
	SRR1188180.fastq.gz	Fastqsanger.gz
	SRR1188182.fastq.gz	Fastqsanger.gz
	SRR1187918.fastq.gz	Fastqsanger.gz
	SRR1188178.fastq.gz	Fastqsanger.gz

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