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Virulence induction in Pseudomonas aeruginosa under inorganic phosphate limitation: a proteomics perspective

ABSTRACT: Inorganic phosphate (Pi) is a central nutrient and signal molecule for bacteria. Pi limitation was shown to increase the virulence of a number of phylogenetically diverse pathogenic bacteria with different in lifestyles. Hypophosphatemia enhances the risk of death in patients due to general bacteremia and was observed after the surgical injury in humans and animals. Phosphate therapy, or the reduction of bacterial virulence by the administration of Pi or phosphate-containing compounds, is a promising anti-infective therapy approach that will not cause cytotoxicity nor the emergence of antibiotic-resistant strains. The proof of concept of phosphate therapy has been obtained using primarily Pseudomonas aeruginosa (PA) as a model. However, a detailed understanding of Pi-induced changes at protein levels is missing. Using pyocyanin production as proxy, we show that the Pi-mediated induction of virulence is a highly cooperative process that occurs between 0.2 to 0.6 mM Pi. We present a proteomics study of PA grown in minimal medium supplemented with either 0.2 or 1 mM Pi and rich medium. About half of the predicted PA proteins could be quantified. Among the 1,471 dysregulated proteins comparing growth in 0.2 mM to 1 mM Pi, 1100 were depleted under Pi-deficient conditions. Most of these proteins are involved in general and energy metabolism, different biosynthetic and catabolic routes, or transport. Pi depletion caused accumulation of proteins that belong to all major families of virulence factor categories, including pyocyanin synthesis, secretion systems, quorum sensing, chemosensory signaling and the secretion of proteases, phospholipases and phosphatases.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Pseudomonas Aeruginosa Pao1

SUBMITTER: Sandra Maass

LAB HEAD: Dörte Becher

PROVIDER: PXD033250 | Pride | 2022-11-02

REPOSITORIES: Pride

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			Action	DRS
	210526_OV3_P5_JB_TKr_2021_016_PAeruginosa_3ul_LB_S1_01.raw	Raw
	210526_OV3_P5_JB_TKr_2021_016_PAeruginosa_3ul_LB_S1_02.raw	Raw
	210526_OV3_P5_JB_TKr_2021_016_PAeruginosa_3ul_LB_S1_03.raw	Raw
	210526_OV3_P5_JB_TKr_2021_016_PAeruginosa_3ul_LB_S1_04.raw	Raw
	210526_OV3_P5_JB_TKr_2021_016_PAeruginosa_3ul_LB_S1_05.raw	Raw

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Project description:Binding of transcription factors to DNA is a key regulatory step in the control of gene expression. DNA sequences with high affinity for transcription factors occur more frequently in the genome than instances of genes bound or regulated by these factors. How specific gene regulation is achieved by transcription factors remains unclear. We used genome-wide approaches to study how trans factors shape the binding and regulatory landscape of Pho4, a budding yeast transcription factor that activates gene expression in response to phosphate limitation. In no phosphate (0mM phosphate) medium, Pho4 is transported into the nucleus and activates transcription of a set of genes (PHO) that are necessary for cell survival in phosphate limited environment. In high phosphate (10mM phosphate) medium, Pho4 is transported outside of the nucleus and the transcription program of PHO genes are turned off. Here we examined the transcription profiling of S. cerevisiae (W303) in various strains treated in media with 10mM or 0mM inorganic phosphate, to study the the transcription activation by Pho4 its cooperative binding factor Pho2, and its competitive binding factor Cbf1. (I) To infer how much Pho4 itself, Pho2 itself, and the interaction between Pho2 and Pho4 contribute to transcriptional activation in response to Pi starvation, we applied mutant cycle analysis to compare wild type, pho2Δ, pho4Δ and pho2Δ pho4Δ strains in both 10 mM and 0 mM Pi conditions. Mutant cycle analysis is a cyclic comparison among all various mutants. For example, by comparing pho2Δ and pho2Δ pho4Δ, we can directly infer the contribution from Pho4 alone to gene activation. And by comparing wild type and pho2Δ, we can infer the contribution from Pho2 alone together with that from the interaction between Pho2 and Pho4. (II) Compare two strains or one strain in two treatments in multiple replicates with dye-swap method. wt no vs high Pi, to identify genes that are induced in response to Pi starvation. cbf1Δ vs wt in high Pi, to identify the influence of Cbf1 on gene expression in high Pi condition. cbf1Δpho4Δ vs wt in high Pi, to identify the influence of Cbf1 on gene expression that is mediated through Pho4, in high Pi condition. rtg3Δtye7Δ vs wt in high Pi, to identify the influence of Rtg3 and Tye7 on gene expression in high Pi condition. pho80Δ vs pho80Δpho4Δ, to identify the genes that are activated Pho4 when Pho4 is fully nuclear localized. pho80Δcbf1Δ vs pho80Δcbf1Δpho4Δ, to identify the influence of Cbf1 on gene expression that is mediated through Pho4, when Pho4 is fully nuclear localized.

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Virulence induction in Pseudomonas aeruginosa under inorganic phosphate limitation: a proteomics perspective

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