Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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A new Rrf2 repressor family member, SaiR, controls transcription of spxA2 encoding a regulator of the oxidative stress response in Bacillus anthracis.

ABSTRACT: Spx, a member of the ArsC (arsenate reductase) protein family, is highly conserved in Gram-positive bacteria, where it interacts with RNA polymerase to activate transcription in response to toxic oxidants.Â In Bacillus anthracis, resistance to oxidative stress requires the activity of two paralogous forms of Spx, SpxA1 and SpxA2.Â Â Suppressor mutations were identified in spxA1 mutant cells that conferred resistance to hydrogen peroxide. Â The mutations, which reside in saiR within the operon that also contains the spxA2 gene, caused derepression of spxA2 transcription. The product of saiR is a member of the Rrf2 family of transcriptional repressors. Â Derepression of spxA2 in a saiR mutant required SpxA2, indicating an autoregulatory mechanism of spxA2 transcriptional activation. Â Reconstruction of SaiR-dependent control of spxA2 was accomplished in Bacillus subtilis, where deletion analysis uncovered two cis-elements within the spxA2 regulatory region that are required for repression.Â Both sites bear sequences of dyad symmetry, mutations in which abolished SaiR binding and SaiR-dependent repression of transcription from the spxA2 promoter in vitro.Â Previous studies have provided evidence that spxA2 is one of the most highly induced genes in a macrophage infected with B. anthracis.Â The work reported herein uncovered a key regulator, SaiR, which controls spxA2 transcriptional activation. Direct comparison of experimental samples to control samples in a two-color hybridization. Two experimental samples were compared against two control samples in triplicate for each.

ORGANISM(S): Bacillus anthracis

SUBMITTER: Bryan Frank

PROVIDER: E-GEOD-57851 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Abstract of associated manuscript: The Bacillus subtilis extracytoplasmic function (ECF) sigma(M) factor is activated by cell envelope stress elicited by antibiotics, and by acid, heat, ethanol and superoxide stresses. Here, we have used several complementary approaches to identify genes controlled by sigma(M). In many cases, expression is only partially dependent on sigma(M) because of both overlapping promoter recognition with other ECF sigma factors and the presence of additional promoter elements. Genes regulated by sigma(M) have a characteristic pattern of induction in response to cell envelope-acting antibiotics as evidenced by hierarchical clustering analysis. sigma(M) also contributes to the expression of the Spx transcription factor and thereby indirectly regulates genes of the Spx regulon. Cell envelope stress responses also include regulons controlled by sigma(W), sigma(B) and several two-component regulatory systems (e.g. LiaRS, YycFG, BceRS). Activation of the sigma(M) regulon increases expression of proteins functioning in transcriptional control, cell wall synthesis and shape determination, cell division, DNA damage monitoring, recombinational repair and detoxification. WT (-van) vs. WT (+van), sigM (-van) vs. sigM (+van), WT (-van) vs. sigM (-van), WT (+van) vs. sigM (+van), WT (-van) vs. spx (-van), WT (+van) vs. spx (+van). Each experiment was conducted at least twice using two independent total RNA preparations. For vancomycin untreated and treated experiments, untreated samples were labeled with Alexa Fluor 555 and treated samples with Alexa Fluor 647. For WT vs. mutant experiments, wild type was labeled with Alexa Fluor 555 and mutants with Alexa Fluor 647. For dye swap experiment, wild-type was labeled with Alexa Fluor 647 and mutant with Alexa Fluor 555. Bacillus subtilis CU1065, WT (-van) vs. WT (+van), sigM (-van) vs. sigM (+van), WT (-van) vs. sigM (-van), WT (+van) vs. sigM (+van), WT (-van) vs. spx (-van), WT (+van) vs. spx (+van)

Project description:Genetic dissection of the S rat genome has provided strong evidence for the presence of two interacting blood pressure (BP) quantitative trait loci (QTLs), termed QTL1 and QTL2, on rat chromosome 5. However, the identities of the underlying interacting genetic factors remain unknown. Further experiments targeted to identify the interacting genetic factors by the substitution mapping approach alone are difficult because of the interdependency of natural recombinations to occur at the two QTLs. We hypothesized that the interacting genetic factors underlying these two QTLs may interact at the level of gene transcription and thereby represent expression QTLs (eQTLs). To detect these interacting eQTLs, a custom QTL chip containing the annotated genes within QTL1 and QTL2 was developed and used to conduct a transcriptional profiling study of S and two congenic strains that retain either one or both the QTLs. The results uncovered an interaction between two transcription factors, DMRTA2 and NFIA. Further, the âbiological signatureâ elicited by these two transcription factors was differential between the congenic strain that retained LEW alleles at both QTL1 and 2 compared to the congenic strain that retained LEW alleles at QTL1 alone. A network of transcription factors potentially affecting BP could be traced, lending support to our hypothesis. Pairs of Cy5 and Cy3 labeled targets were co-hybridized onto either a custom long oligonucleotide microarray for the interrogation of 231 genes encompassed by QTL1 and QTL2, or a TIGR rat cDNA array consisting of 26,401 probe elements representing 20,465 unique non-QTL genes. A âflip-dyeâ or âbalanced blockâ design was used as the experimental method of choice to account for potential dye-bias labeling effects. Six âbalanced blockâ normalized files are submitted for the long oligonucleotide array interrogating the hearts from S versus S.LEW(5)x6x9 animals, fourteen âflip-dyeâ hybridizations are submitted for the cDNA array interrogating the hearts from S versus S.LEW(5)x6x9 animals, and twelve âflip-dyeâ hybridizations are submitted for the cDNA array interrogating the hearts from S versus S.LEW(5)x6x11 animals. GPR and MEV files cannot be located.

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A new Rrf2 repressor family member, SaiR, controls transcription of spxA2 encoding a regulator of the oxidative stress response in Bacillus anthracis.

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