Project description:To identify the transcriptional targets of the DNA-binding response regulator HnoC (SO_2540), mRNA transcript levels in Shewanella oneidensis were measured using whole genome microarray analysis. Transcript levels were compared between WT Shewanella oneidensis and a hnoC deletion strain.
Project description:Silencing pathways prevent transposable element (TE) proliferation and help to maintain genome integrity through cell division. Silenced genomic regions can be classified as either euchromatic or heterochromatic, and are targeted by genetically separable epigenetic pathways. In plants, the RNA-directed DNA methylation (RdDM) pathway targets mostly euchromatic regions, while CMT methyltransferases are mainly associated with heterochromatin. However, many epigenetic features - including DNA methylation patterning - are largely indistinguishable between these regions, so how the functional separation is maintained is unclear. The linker histone H1 is preferentially localised to heterochromatin and has been proposed to restrict RdDM from encroachment. To test this hypothesis, we followed RdDM genomic localisation in an H1 mutant by performing ChIP-seq on the largest subunit, NRPE1, of the central RdDM polymerase (Pol V). Loss of H1 resulted in heterochromatic TE enrichment of NPRE1. Increased NRPE1 binding was associated with increased chromatin accessibility in h1, suggesting that H1 restricts NRPE1 occupancy by compacting chromatin. However, RdDM occupancy did not impact H1 localization, demonstrating that H1 hierarchically restricts RdDM positioning. H1 mutants experience major symmetric (CG and CHG) DNA methylation gains, and by generating an h1/nrpe1 double mutant, we demonstrate these gains are largely independent of RdDM. However, loss of NRPE1 occupancy from a subset of euchromatic regions in h1 experienced corresponding loss of methylation in all sequence contexts, while at ectopically bound heterochromatic loci, NRPE1 deposited methylation specifically in the CHH context. Additionally, we found that H1 restricts the occupancy of the methylation reader and activator complex component, SUVH1, indicating that H1’s regulatory control of methylation pathways is not limited to RdDM. Together, the results support a model whereby H1 helps maintain the exclusivity of heterochromatin by preventing encroachment from other competing pathways.
Project description:Comparisson of expression profiling of a etrA deletion mutant strain (experimental sample) with that of the wild type Shewanella oneidensis MR-1 strain to assess global direct/indirect genetic regulation EtrA in Shewanella oneidensis MR-1 shares 73.6% and 50.8% amino acid sequence identity with the oxygen-sensing regulator Fnr in E. coli and Anr in Pseudomonas aeruginosa, respectively; however, its regulatory role of anaerobic metabolism in Shewanella spp. is complex and not well understood. Whole-genome expression profiling using a etrA gene deletion mutant as the experimental sample and the wild type strain as the reference, determine that EtrA fine-tunes the expression of genes involved in various anaerobic metabolic pathways, including nitrate, fumarate and dimethyl sulfoxide reduction. Moreover, genes involved in prophage activation and and genes implicated in aerobic metabolism were also differentially expressed. In contrast to previous studies that attributed a minor regulatory role to EtrA in Shewanella spp., this study demonstrates that EtrA acts as a global transcriptional regulator and cofers physiological advantages to the strain under certain growth conditions.
Project description:To identify the transcriptional targets of the DNA-binding response regulator HnoC (SO_2540), mRNA transcript levels in Shewanella oneidensis were measured using whole genome microarray analysis. Transcript levels were compared between WT Shewanella oneidensis and a hnoC deletion strain. Transcript levels of a WT and hnoC deletion strain were measured after 15 hrs growth, 4 independent replicates were performed for each strain
