Unknown,Transcriptomics,Genomics,Proteomics

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M.ruminantium_B.proteoclasticus_coculture_vs_monocultures

ABSTRACT: This study was undertaken with two major goals in mind: i) To investigate the potential for interspecies hydrogen transfer between a hemicellulytic rumen bacterium (B. proteoclasticus) and a methanogenic achaea (M. ruminantium) Microscopic examination had previously shown B. proteoclasticus in co-culture with M. ruminantium rapidly formed cell to cell co-aggregates. ii) To examine the expression of genes involved in methanogenesis under more rumen-like conditions. Cells of M.ruminantium and B. proteoclasticus were grown in triplicate as mono- or co-cultures until a mid-exponential growth phase was reached. RNAs were extracted and purified. The relative quantities of RNAs contributed by each organism to the co-culture samples were determined by rt qPCR using genes previously shown to be constitutively expressed. The mono-culture RNAs were then combined in the determined proportions to normalise mRNA abundance with their co-culture replicates. cDNAs were synthesized and labelled and hybridised to the arrays. 3 biological replicates were used and the cDNA from each was divided in two to allow each to undergo a dyeswap (total of six arrays) giving technical replicates. Each gene was probed in triplicate adding further technical replication. Blanks and negative controls were also included. The log ratios for each probes replicates (biological and technical) were averaged prior to further analysis.

ORGANISM(S): Butyrivibrio proteoclasticus B316

SUBMITTER: Carl Yeoman

PROVIDER: E-GEOD-18716 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Candida albicans and Candida dubliniensis are closely related species displaying differences in virulence and genome content, therefore providing potential opportunities to identify novel C. albicans virulence genes. C. albicans gene arrays were used for comparative analysis of global gene expression in the two species in reconstituted human oral epithelium (RHE). C. albicans (SC5314) showed upregulation of hypha-specific and virulence genes within 30 min postinoculation, coinciding with rapid induction of filamentation and increased RHE damage. C. dubliniensis (CD36) showed no detectable upregulation of hypha-specific genes, grew as yeast, and caused limited RHE damage. Several genes absent or highly divergent in C. dubliniensis were upregulated in C. albicans. One such gene, SFL2 (orf19.3969), encoding a putative heat shock factor, was deleted in C. albicans. ÎÎsfl2 cells failed to filament under a range of hypha-inducing conditions and exhibited greatly reduced RHE damage, reversed by reintroduction of SFL2 into the ÎÎsfl2 strain. Moreover, SFL2 overexpression in C. albicans triggered hyphal morphogenesis. Although SFL2 deletion had no apparent effect on host survival in the murine model of systemic infection, ÎÎsfl2 strain-infected kidney tissues contained only yeast cells. These results suggest a role for SFL2 in morphogenesis and an indirect role in C. albicans pathogenesis in epithelial tissues. Global gene expression in Candida albicans SC5314 or Candida dubliniensis CD36 on reconstituted human oral epithelium (RHE) 30 min postinoculation. Gene expression on RHE was compared to gene expression in cells used as inoculum (0 min). Performed in three biological replicates with dye swaps across the biological replicates. Gene expression of Candida cells on RHE was normalized to gene expression in reference control (0 min); log2 ratios were calculated by dividing spot intensity of experimental by that of the reference control.

Project description:Most homeodomains are unique within a genome, yet many are highly conserved across vast evolutionary distances, implying strong selection on their precise DNA-binding specificities. We determined the binding preferences of the majority (168) of mouse homeodomains to all possible 8-base sequences, revealing rich and complex patterns of sequence specificity, and showing for the first time that there are at least 65 distinct homeodomain DNA-binding activities. We developed a computational system that successfully predicts binding sites for homeodomain proteins as distant from mouse as Drosophila and C. elegans, and we infer full 8-mer binding profiles for the majority of known animal homeodomains. Our results provide an unprecedented level of resolution in the analysis of this simple domain structure and suggest that variation in sequence recognition may be a factor in its functional diversity and evolutionary success. Keywords: Mouse homeodomain protein binding microarrays 178 Protein binding microarray (PBM) experiments of mouse homeodomains were performed, with 10 proteins done in replicate. Briefly, the PBMs involved binding GST-tagged mouse homeodomains to custom-designed, double-stranded 44K Agilent microarrays in order to determine their sequence preferences. The method is described in Berger et al., Nature Biotechnology 2006. A key feature is that the microarrays are composed of de Bruijn sequences that contain each 10-base sequence once and only once, providing an evenly balanced sequence distribution. Individual de Bruijn sequences have different properties, including representation of gapped patterns. The array sequences as well as the primary array data are available via a EULA at http://the_brain.bwh.harvard.edu/pbms/webworks2/. Here we provide the data transformed into median intensities (after normalization and detrending of the original array data) for all 32,896 8-base sequences, Z-scores for these intensities, and E-scores. E-scores are a modified version of AUC, and describe how well each 8-mer ranks the intensities of the spots. In general the E-scores are slightly more reproducible than Z-scores, but contain less information about relative binding affinity. Additional experimental details are found in Berger et al., Nature Biotechnology 2006, Berger et al., Cell 2008, and the accompanying Supplementary information.

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M.ruminantium_B.proteoclasticus_coculture_vs_monocultures

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