Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcriptional analysis of prebiotic uptake and catabolism by Lactobacillus acidophilus NCFM

ABSTRACT: The global transcriptome of the wild type Lactobacillus acidophilus NCFM strain (NCK56) was measured during exponential growth on 11 prebiotic carbohydrates and glucose to identify the specific gene cluster differentially upregulated in response to each carbohydrate. Oligoarray hybridization experiments were performed to compare the differential transcriptional profiles of the NCK56 at the early-log (OD600nm 0.3-0.5) phase when cultured in semi-synthetic medium (SSM, Barrangou et al., 2003 PNAS. 100:8957-8962, supplemented with a range of 1% (w/v) carbohydrates, at 37 M-BM-0C and harvested by centrifugation and flash freezing. A total of 12 NCK56 cultures were prepared by growing in SSM supplemented with each one of the 12 tested carbohydrates. Each culture was grown in fresh carbohydrate supplemented SSM with a 1% inoculum from an overnight culture for a total of five subcultured passages. Aliquots of cells were collected at OD600nm of 0.3-0.5 (early log-phase) for total RNA extraction and cDNA synthesis. Labeled cDNA samples from NCK56 were coupled with mono-reactive Cy3 and Cy5 dyes (GE Healthcare Bio-Sciences Corp., Piscataway, NJ), for two technical replicates on each carbohydrate (e.g. cy3-GOS and cy5-GOS). Comparative hybridizations were performed on samples representing two different carbohydrate cultures at same growth phases labeled with either cy3 or cy5 using a loop design for 12 hybridizations.

ORGANISM(S): Lactobacillus acidophilus NCFM

SUBMITTER: Todd Klaenhammer

PROVIDER: E-GEOD-35968 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Publications

Transcriptional analysis of prebiotic uptake and catabolism by Lactobacillus acidophilus NCFM.

Andersen Joakim Mark JM Barrangou Rodolphe R Hachem Maher Abou MA Lahtinen Sampo J SJ Goh Yong-Jun YJ Svensson Birte B Klaenhammer Todd R TR

PloS one 20120919 9

The human gastrointestinal tract can be positively modulated by dietary supplementation of probiotic bacteria in combination with prebiotic carbohydrates. Here differential transcriptomics and functional genomics were used to identify genes in Lactobacillus acidophilus NCFM involved in the uptake and catabolism of 11 potential prebiotic compounds consisting of α- and β-linked galactosides and glucosides. These oligosaccharides induced genes encoding phosphoenolpyruvate-dependent sugar phosphotra ...[more]

PMID: 23028535

Similar Datasets

Project description:The complexity of the plant cell walls creates many challenges for microbial decomposition. Clostridium phytofermentans, an anaerobic bacterium isolated from forest soil, directly breaks down and utilizes many plant cell wall carbohydrates. The objective of this research is to understand constraints on rates of plant decomposition by C. phytofermentans and identify molecular mechanisms that may overcome these limitations. Experimental evolution via repeated serial transfers during exponential growth was used to select for C. phytofermentans genotypes that grow more rapidly on cellobiose, cellulose and xylan. To identify the underlying mutations an average of 13,600,000 paired-end 100bp paired end reads were generated per population resulting in ~300 fold coverage of each site in the genome. A mixture of alleles fixed in the population down to frequencies of 5% could be identified with statistical confidence. Many mutations are in carbohydrate-related genes including the promoter regions of glycoside hydrolases and amino acid substitutions in ABC transport proteins involved in carbohydrate uptake, signal transduction sensors that detect specific carbohydrates, proteins that affect the export of extracellular enzymes, and regulators of unknown specificity. Protein structural modeling of the ABC transporter complex proteins suggests that mutations in these genes may alter the recognition of carbohydrates by substrate-binding proteins and communication between the intercellular face of the transmembrane proteins and the ATPase binding protein. Experimental evolution was effective in identifying molecular constraints on the rate of cellulose and hemicellulose fermentation and selected for putative gain of function mutations that do not typically appear in traditional molecular genetic screens. The results reveal new strategies for evolving and engineering microorganisms for faster growth on plant carbohydrates. Experimental evolution via repeated serial transfers during exponential growth was used to select for C. phytofermentans genotypes that grow more rapidly on cellobiose. Microarray analysis was performed on 1 time point from the evolved lines and the initial founder

Project description:Genomic analysis of the model lignocellulosic biomass degrading bacteria C. phytofermentans indicates that it can degrade, transport, and utilize a wide-range of carbohydrates as possible growth substrates. Previous experiments characterized the expression of the degradation and transport machinery using custom whole genome oligonucleotide microarrays. The results indicate that C. phytofermentans utilizes ATP-binding cassette (ABC) transporters for carbohydrate uptake and does not use the sole phosphoenolpyruvate-phosphotransferase system (PTS) for any of the tested substrates. While some ABC transporters are specific for a single carbohydrate, the expression profiles indicate that others may be capable of transporting multiple substrates. Distinct sets of Carbohydrate Active Enzymes (CAZy) genes were also up-regulated on specific substrates indicative of C. phytofermentans ability to selectively degrade plant biomass. We also identified a highly expressed cluster of genes which includes seven extracellular glycoside hydrolases and two ABC transporters with unknown specificity. These results lead to the hypothesis that when grown on plant biomass, C. phytofermentans is capable of degrading and transporting all major carbohydrate components of the plant cell. To test this, C. phytofermentans was grown on cornstover and switchgrass. Results from this expression data and HPLC analysis indicates that C. phytofermentans is utilizing multiple substrates. with multiple sugar ABC transporter clusters and glycoside hydrolases being expressed. Interestingly all of the transporters were initially identified on disaccharides or oligio-saccharides, and none of the transporters identified as monosaccharide specific transporters were expressed. This could be an indication that C. phytofermentans prefers to transport oligiosacchrides over monosaccharides. The results presented here corroborate the genomic data which indicates the breath of the carbohydrate degradation, transport, and utilization machinery of C. phytofermentans. C. phytofermentans was cultured anaerobically on switchgrass and corn stover to determine specific expression patterns. The data in this series consists three independent RNA preparations from replicate cultures.

Dataset Information

Transcriptional analysis of prebiotic uptake and catabolism by Lactobacillus acidophilus NCFM

Publications

Transcriptional analysis of prebiotic uptake and catabolism by Lactobacillus acidophilus NCFM.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure