Project description:Metaproteomic analysis of an enriched anaerobic rumen consortium (ERAC) using sugarcane bagasse and rumen as unique carbon and microbial sources
Project description:Background and aims The endophytic diazotrophic strain CBAmC of Nitrospirillum amazonense has been reported as a plant growth promoter of sugarcane variety RB867515 when grown under field conditions. The present work aimed to assess the influence of apoplast fluid from RB867515 on the transcriptomic and proteomic profiles of CBAmC cultured in vitro. Methods RNA-Seq in Ion Proton™ and ESI-LC-MS/MS peptide analysis were used to evaluate the transcriptomic and proteomic profiles, respectively, of CBAmC exposed for 2 h to the sugarcane apoplast fluid. Results The bacterial transcriptomic and proteomic profiles were well correlated. The overall response of CBAmC to the apoplast fluid included overexpression of defense systems against reactive oxygen species (ROS) and osmotic stress, RND efflux pumps for toxic compounds, Sec and Tat secretory systems, and assimilative metabolism of iron. In contrast, active transporters of organic compounds, chemotaxis system and flagellum structure were underexpressed. Conclusions The bacterial metabolic pathways / functions activated in response to the sugarcane apoplast fluid are most likely related to its adaptation to the peculiar characteristics of the fluid. The activation of some of those functions could be determinant for its adaptation to the sugarcane apoplastic niche, and perhaps be involved in the previously observed effect of promoting plant growth. SUBMITTER_CITATION: Terra, L.A., de Soares, C.P., Meneses, C.H.S.G. et al. Plant Soil (2019). Transcriptome and proteome profiles of the diazotroph Nitrospirillum amazonense strain CBAmC in response to the sugarcane apoplast fluid.
Project description:Background: Wood-decay basidiomycetes are effective for the degradation of highly lignified and recalcitrant substrates. Brown-rot strain produces carbohydrate-active enzymes involved in the degradation of lignocellulosic materials, along with a non-enzymatic mechanism, via Fenton reaction. Differences in the lignocellulose metabolism occurring even among closely related brown-rots are not completely understood, bringing attention to a multi-omics study of brown-rot L. sulphureus. Results: To evidence the oxidative-hydrolytic mechanism, the Laetiporus sulphureus ATCC 52600 genome was sequenced and the response to lignocellulosic substrates was analyzed by transcriptomics and proteomics. The transcriptomic profile in response to a short cultivation period on in natura sugarcane bagasse revealed 128 out of 12,802 upregulated transcripts. The high upregulated transcripts included a set of redox enzymes along with hemicellulases. The exoproteome in response to extended-time cultivation with Avicel, and steam-exploded sugarcane bagasse, sugarcane straw, and Eucalyptus grandis revealed 121 proteins. Contrasting to the mainly oxidative profile observed in the transcriptome, the secretomes showed a diverse hydrolytic repertoire including constitutive cellulases and hemicellulases, in addition to 19 proteins upregulated relative to glucose. The secretome produced on sugarcane bagasse was evaluated in the saccharification of pretreated sugarcane straw by supplementing a commercial cocktail. Additionally, growth analysis revealed that L. sulphureus ATCC 52600 has higher efficiency to assimilate glucose than other mono and disaccharides. Conclusion: This study shows the singularity of L. sulphureus ATCC 52600 relative to other Polyporales brown-rots, regarding the presence of cellobiohydrolase and peroxidase class II. The multi-omic analysis reinforces the oxidative-hydrolytic metabolism involved in lignocellulose deconstruction, providing insights into the overall mechanisms as well as specific proteins of each step.
Project description:Transcriptional profiling of A. niger comparing WT strain vs. ΔXlnR strain treated with steam-exploded sugarcane bagasse (SESB) for 6, 12 and 24 h. The main objective was to identifiy genes related to cellulases and hemicellulases, comparing the differences between WT strain and the strain with the disrupted xylanolytic transcriptional activator gene, XlnR, after treatment with steam-exploded sugarcane. The experiment was further validated by real-time PCR, mass spectrometry of secreted proteins and enzymatic assays.
Project description:Transcriptional profiling of A. niger comparing WT strain vs. ÎXlnR strain treated with steam-exploded sugarcane bagasse (SESB) for 6, 12 and 24 h. The main objective was to identifiy genes related to cellulases and hemicellulases, comparing the differences between WT strain and the strain with the disrupted xylanolytic transcriptional activator gene, XlnR, after treatment with steam-exploded sugarcane. The experiment was further validated by real-time PCR, mass spectrometry of secreted proteins and enzymatic assays. Three-condition experiment : WT-SESB or ÎXlnR-SESB for 6, 12 and 24 h at 30 oC in batch culture. Firstly, WT and ÎXlnR strains were grown in minimal medium with fructose as carbon source (control), and then transferred to SESB as carbon source.
Project description:Raw LC-MSMS data from the Plos One publication: "Novel Reusable Animal Model for Comparative Evaluation of In vivo Growth and Protein-Expression of Escherichia coli O157 strains in the Bovine Rumen" Represents 2 LC-MSMS runs L (lactation diet) and M (maintenance diet) in 10 fractions each. Each run contains 6 iTRAQ labeled samples, 3 strains grown in both the in-vivo and in-vitro rumen fluid.
Project description:The transcriptome of Escherichia coli K-12 has been widely studied over a variety of conditions for the past decade while such studies involving E. coli O157:H7, its pathogenic cousin, are just now being conducted. To better understand the impact of rumen fluid on E. coli O157:H7, global transcript levels of strain EDL933 cells resuspended in heat clarified rumen fluid for 15 min were compared to cells resuspended in fresh LB using microarrays. Seven independent RNA samples from rumen fluid treated cultures were paired with seven independent RNA samples from control cultures for hybridization to seven two-color microarrays. For three arrays, the control RNA sample was labeled with Cy3 dye and the experimental RNA sample was labeled with Cy5 dye, the dyes were reversed for the other four arrays to overcome dye bias.
Project description:The phytopathogenic fungus Chrysoporthe cubensis is a relevant source of lignocellulolytic enzymes. This work aimed to compare the profile of lignocellulose- degrading proteins secreted by C. cubensis grown under semi-solid state fermentation using wheat bran and sugarcane bagasse. The proteins from the fungus extract grown in wheat bran (WBE) and sugarcane bagasse (SBE) were qualitative and quantitatively analyzed by liquid chromatography-electrospray ionization tandem mass spectrometry (LC–ESI–MS/MS). Label-free proteomic analysis of WBE and SBE showed that the fungus produced a spectrum of carbohydrate-active enzymes (CAZymes) with exclusive characteristics from each extract. While SBE resulted in an enzymatic profile directed towards the depolymerization of cellulose, the enzymes in WBE were more adaptable to the degradation of biomass rich in hemicellulose and other non-lignocellulosic polymers. Saccharification of alkaline pre-treated sugarcane bagasse with SBE promoted glucose release higher than commercial cocktails (8.11 g L -1 ), while WBE promoted the higher release of xylose (5.71 g L -1 ). Our results allowed an in-depth knowledge of the complex set of enzymes secreted by C. cubensis responsible for its high lignocellulolytic activity and still provided the identification of promising target proteins for biotechnological applications in the context of biorefinery.