Dataset Information

Inorganic Phosphate Limitation Modulates Capsular Polysaccharide Composition in Mycobacteria.

ABSTRACT: Mycobacterium tuberculosis is protected by an unusual and highly impermeable cell envelope that is critically important for the successful colonization of the host. The outermost surface of this cell envelope is formed by capsular polysaccharides that play an important role in modulating the initial interactions once the bacillus enters the body. Although the bioenzymatic steps involved in the production of the capsular polysaccharides are emerging, information regarding the ability of the bacterium to modulate the composition of the capsule is still unknown. Here, we study the mechanisms involved in regulation of mycobacterial capsule biosynthesis using a high throughput screen for gene products involved in capsular ?-glucan production. Utilizing this approach we identified a group of mutants that all carried mutations in the ATP-binding cassette phosphate transport locus pst These mutants collectively exhibited a strong overproduction of capsular polysaccharides, including ?-glucan and arabinomannan, suggestive of a role for inorganic phosphate (Pi) metabolism in modulating capsular polysaccharide production. These findings were corroborated by the observation that growth under low Pi conditions as well as chemical activation of the stringent response induces capsule production in a number of mycobacterial species. This induction is, in part, dependent on ? factor E. Finally, we show that Mycobacterium marinum, a model organism for M. tuberculosis, encounters Pi stress during infection, which shows the relevance of our findings in vivo.

SUBMITTER: van de Weerd R

PROVIDER: S-EPMC4882446 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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Inorganic Phosphate Limitation Modulates Capsular Polysaccharide Composition in Mycobacteria.

van de Weerd Robert R Boot Maikel M Maaskant Janneke J Sparrius Marion M Verboom Theo T van Leeuwen Lisanne M LM Burggraaf Maroeska J MJ Paauw Nanne J NJ Dainese Elisa E Manganelli Riccardo R Bitter Wilbert W Appelmelk Ben J BJ Geurtsen Jeroen J

The Journal of biological chemistry 20160404 22

Mycobacterium tuberculosis is protected by an unusual and highly impermeable cell envelope that is critically important for the successful colonization of the host. The outermost surface of this cell envelope is formed by capsular polysaccharides that play an important role in modulating the initial interactions once the bacillus enters the body. Although the bioenzymatic steps involved in the production of the capsular polysaccharides are emerging, information regarding the ability of the bacte ...[more]

PMID: 27044743

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Project description:BACKGROUND: Bacterial cell surface-associated polysaccharides are involved in the interactions of bacteria with their environment and play an important role in the communication between pathogenic bacteria and their host organisms. Cell surface polysaccharides of probiotic species are far less well described. Therefore, improved knowledge on these molecules is potentially of great importance to understand the strain-specific and proposed beneficial modes of probiotic action. RESULTS: The Lactobacillus plantarum WCFS1 genome encodes 4 clusters of genes that are associated with surface polysaccharide production. Two of these clusters appear to encode all functions required for capsular polysaccharide formation (cps2A-J and cps4A-J), while the remaining clusters are predicted to lack genes encoding chain-length control functions and a priming glycosyl-transferase (cps1A-I and cps3A-J). We constructed L. plantarum WCFS1 gene deletion mutants that lack individual (?cps1A-I, ?cps2A-J, ?cps3A-J and ?cps4A-J) or combinations of cps clusters (?cps1A-3J and ?cps1A-3I, ?cps4A-J) and assessed the genome wide impact of these mutations by transcriptome analysis. The cps cluster deletions influenced the expression of variable gene sets in the individual cps cluster mutants, but also considerable numbers of up- and down-regulated genes were shared between mutants in cps cluster 1 and 2, as well as between mutant in cps clusters 3 and 4. Additionally, the composition of overall cell surface polysaccharide fractions was altered in each mutant strain, implying that despite the apparent incompleteness of cps1A-I and cps3A-J, all clusters are active and functional in L. plantarum. The ?cps1A-I strain produced surface polysaccharides in equal amounts as compared to the wild-type strain, while the polysaccharides were characterized by a reduced molar mass and the lack of rhamnose. The mutants that lacked functional copies of cps2A-J, cps3A-J or cps4A-J produced decreased levels of surface polysaccharides, whereas the molar mass and the composition of polysaccharides was not affected by these cluster mutations. In the quadruple mutant, the amount of surface polysaccharides was strongly reduced. The impact of the cps cluster mutations on toll-like receptor (TLR)-mediated human nuclear factor (NF)-?B activation in host cells was evaluated using a TLR2 reporter cell line. In comparison to a L. plantarum wild-type derivative, TLR2 activation remained unaffected by the ?cps1A-I and ?cps3A-J mutants but appeared slightly increased after stimulation with the ?cps2A-J and ?cps4A-J mutants, while the ?cps1A-3J and ?cps1A-3J, ?cps4A-J mutants elicited the strongest responses and clearly displayed enhanced TLR2 signaling. CONCLUSIONS: Our study reveals that modulation of surface glycan characteristics in L. plantarum highlights the role of these molecules in shielding of cell envelope embedded host receptor ligands. Although the apparently complete cps clusters (cps2A-J and cps4A-J) contributed individually to this shielding, the removal of all cps clusters led to the strongest signaling enhancement. Our findings provide new insights into cell surface glycan biosynthesis in L. plantarum, which bears relevance in the context of host-cell signaling by probiotic bacteria.

Dataset Information

Inorganic Phosphate Limitation Modulates Capsular Polysaccharide Composition in Mycobacteria.

Publications

Inorganic Phosphate Limitation Modulates Capsular Polysaccharide Composition in Mycobacteria.

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