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EslB suppressor screen

ABSTRACT: Suppressors of an Listeria monocytogenes eslB mutant were selected on non-permissive growth conditions

PROVIDER: PRJEB55822 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR10162252_1.fastq.gz	Fastqsanger.gz
	ERR10162252_2.fastq.gz	Fastqsanger.gz
	ERR10162253_1.fastq.gz	Fastqsanger.gz
	ERR10162253_2.fastq.gz	Fastqsanger.gz
	ERR10162254_1.fastq.gz	Fastqsanger.gz

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Project description:In several gram-positive bacterial genera including Bacillus, Staphylococcus, and Listeria, sigma B (σB) has been identified as a stress-responsive alternative sigma factor responsible for initiating transcription of genes (the σB regulon) involved in response to stress-inducing environmental conditions. In L. monocytogenes, a foodborne pathogen of considerable threat to public health and the food industry, σB is involved in regulation of stress response and virulence gene expression. We have defined the σB regulon in L. monocytogenes during early stationary phase and under salt stress (0.3M NaCl) conditions using whole-genome microarrays, identifying 168 genes that generated ≥2.0-fold higher signals in the parental strain 10403S than in an isogenic sigB null mutant (ΔsigB), categorized into nine functional groups including stress-response genes (12), virulence genes (5), and genes related to transport (26) and metabolism (45). To gain a broader biological perspective of the σB stress response system, we applied these microarrays to Listeria innocua under the same environmental conditions. Our studies revealed 64 candidates in the L. innocua σB regulon with ≥2.0-fold higher signals in the parent than in a ΔsigB mutant; 49 of the 64 genes overlap with the L. monocytogenes σB regulon, indicating extensive overlap in σB-controlled genes between the two species. Further transcriptional analysis using TaqMan quantitative real time RT-PCR was performed for selected genes that displayed contrasting fold changes among the four microarray data sets (two stress conditions per species). We report novel members of the L. monocytogenes σB regulon, as well as the initial definition of the L. innocua σB regulon. Our comparative studies of the σB stress response systems in L. monocytogenes and L. innocua revealed features of the σB regulon that are conserved and unique to the two species. Keywords: Listeria monocytogenes, Listeria innocua, SigB regulon, salt stress, stationary phase

Project description:PrfA activity was studied in L. monocytogenes strain EGD and in an isogenic prfA deletion mutant (EGDΔprfA) carrying multiple copies of the wild-type prfA or the mutant prfA* gene (strains EGDΔprfApPrfA and EGDΔprfApPrfA*) after growth in brain heart infusion (BHI), Luria-Bertani broth (LB) or a defined minimal medium (MM) supplemented either with one of the three PTS-carbohydrates, glucose, mannose and cellobiose, or the non-PTS carbon source glycerol. Low PrfA activity was observed in the wild-type EGD strain in BHI and LB with either of these carbon sources, while PrfA activity was high in minimal medium in presence of glycerol but significantly reduced in presence of cellobiose. The strains expressing the prfA and prfA* gene under the prfA promoters, P1 and P2, produced equally large amounts of PrfA protein and high PrfA activity was observed in strain EGDΔprfApPrfA* under all growth conditions. In contrast, high PrfA activity in strain EGDΔprfApPrfA was only observed when this strain was cultured in BHI but not in LB or MM (in presence of either carbon source). A ptsH mutant (lacking a functional HPr) was able to grow in BHI suggesting that growth of L. monocytogenes in this culture medium is supported by carbon sources whose uptake and metabolism are independent of the PTS pathway. However, this mutant was unable to grow in LB and MM regardless which of the four carbon sources was added, suggesting that uptake of the used carbohydrates and the catabolism of glycerol depend fully on the functional common PTS pathway. Furthermore, the growth rates of L. monocytogenes are strongly reduced in presence of large amounts of PrfA protein when growing MM but less in LB and only slightly in BHI. The expression profiles of the genes encoding PTS permeases were determined in the three strains under various growth conditions. The data suggest that PrfA activity correlates with the expression level and the phosphorylation state of specific PTS permeases. This SuperSeries is composed of the following subset Series: GSE12143: Listeria monocytogenes EGD after growth BHI vs. LB vs. MM GSE12145: Listeria monocytogenes EGDΔprfApPrfA and EGDΔprfApPrfA* compared to the wild type strain EGD GSE12146: Listeria monocytogenes EGD and EGD-e

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EslB suppressor screen

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