Gene Co-Expression Network Analysis in Rhodobacter capsulatus and Application to Comparative Expression Analysis of Rhodobacter sphaeroides
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ABSTRACT: Transcriptomic data has been previously used to determine expression patterns in various R. capsulatus strains using custom made Affymetrix microarrays. Additional expression analyses have since been carried out to obtain preliminary data on certain wild type and mutant strains that have not been reported on. We used all current microarray expression data available and constructed an R. capsulatus gene co-expression network and performed functional analysis of identified gene modules. RNA was harvested from various wild type and mutant R. capsulatus strains grown under photoheterotrophic conditions to different growth phases. The samples were hybridized to custom made R. capsulatus Affymetrix microarrays according to manufacturers recommendations. The raw data was RMA normalized and log transformed.
Project description:Rhodobacter capsulatus produces a gene transfer agent (GTA) called RcGTA. RcGTA is a phage-like particle that packages R. capsulatus DNA and transfers it to other R. capsulatus cells. Low-resolution techniques suggested RcGTA packages random DNA. Analysis of the RcGTA terminase sequence revealed a distant relationship to the phage T4 terminase protein, which can also perform sequence-independent packaging. We quantified the relative frequency of packaging for each gene in the genome by hybridization of DNA from RcGTA particles to an R. capsulatus microarray. All genes were found within the RcGTA particles, and random packaging was observed at a genome-wide scale. However, the genes encoding the RcGTA particle were under-packaged compared to other regions. Single-cell expression analysis demonstrated that RcGTA gene expression is not uniform within a culture. We propose a mechanism by which high levels of RcGTA gene transcription in the most active RcGTA producing cells hinders access of the packaging machinery to these genes, which causes a reduction in their packaging frequency. This sub-population gene expression phenomenon likely explains the lack of observed cell lysis in RcGTA producing cultures. DNA extracted from within RcGTA particles produced by DE442, an RcGTA overproducer, was hybridized to an Affymetrix genechip custom array in order to quantify the relative frequency with which any given gene was packaged within the particles. These data were MAS5 normalized. An underlying packaging bias was suspected to be transcription related, so these frequencies were compared to expression levels within the population, as determined by hybridization of late-stationary RNA samples of DE442 and of the wildtype (SB1003) to the same custom array. These data were RMA normalized.
Project description:Expression profiles of wild type and gtaI mutant in logarithmic and stationary phase of growth. The objective was to search for differentially regulated genes potentially associated with EPS production in R. capsulatus. A single sample of each strain in both growth phases for basic comparison of expression profiles.
Project description:Transcriptomic data has been previously used to determine expression patterns in various R. capsulatus strains using custom made Affymetrix microarrays. Additional expression analyses have since been carried out to obtain preliminary data on certain wild type and mutant strains that have not been reported on. We used all current microarray expression data available and constructed an R. capsulatus gene co-expression network and performed functional analysis of identified gene modules.
Project description:The diazotrophic bacterium Rhodobacter capsulatus synthesizes a molybdenum nitrogenase and an alternative iron-only nitrogenase, enabling growth with molecular dinitrogen as sole nitrogen source. Regulation of nitrogen fixation was analyzed by proteome profiling of wild-type and mutant strains lacking the transcriptional regulators NifA, AnfA, and MopAB.
Project description:The samples in this series were used to analyze the transcriptome of the CtrA regulon using wild type (SB1003) and ctrA mutant (SBRM1) strains of Rhodobacter capsulatus. As well, the transcriptome of growth phase regulation in R. capsulatus SB1003 between log and stationary phases was determined.
Project description:Global transcriptome analyses at growth before and after 10 min of photooxidative stress were applied to monitor stress dependent gene expression in the alpha-proteobacterium Rhodobacter capsulatus. Transcriptome profiles of pigmented cultures with high aeration were monitored before and after the onset of singlet oxygen stress.
Project description:The diazotrophic bacterium Rhodobacter capsulatus is able to synthesize two nitrogenases, a molybdenum-dependent and an alternative Mo-free iron-only nitrogenase, enabling growth with molecular dinitrogen (N2) as sole nitrogen source. The Mo response of the wild type and a mutant lacking the high-affinity molybdate transporter, ModABC, were analyzed by proteome profiling.