Team:Colombia/Project/Experiments/Aliivibrio and Streptomyces

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Aim

Our system heavily relies in the ability of our bacteria to sense environmental cues belonging to the pathogens we’re particularly concerned of. Chitin is one of the main components of fungal cell walls and, in consequence, we will use it as an indicator of fungal infection.

Figure 2. Model for the regulation of the chitinolytic system in S. coelicolor. (a) Repression and (b) induction of the chitinolytic system in S. coelicolor. In the presence of chitobiose or a related metabolic product of chitin, ChiS is autophosphorylated at a conserved histidine residue (H1199). The phosphoryl group is then transferred to a conserved aspartate (D54) of ChiR, which then binds to the chi promoter regions to activate expression of chitinolytic (chi) genes. The repression of chi genes, would be the result of binding of DasA to the extracellular domain of the sensor ChiS, ‘locking’ it into an inactive conformation. In the presence of chitin, the extracellular signal, typically (GlcNAc)2, competes with the sensor ChiS for binding to DasA. ChiS could not bind the DasA–(GlcNAc)2 complex and would thereby be activated to the ‘plus’ phenotype, followed by the transcriptional activation of the chitinolytic genes by ChiR. Taken from: Colson et al., 2003

Figure 1. A: Negative phenotype for chitin sensing. B:Dimers of N-acetylglucosamine (GlcNAc)2 pass through the outer membrane (OM) helped by a chitoporin and bind a chitin-binding protein (CBP), which unlocks the histidine kinase sensor thus starting the signalling. This leads to the activation of chitin metabolic genes. Taken from: Li and Roseman, 2003

Vibrio fischeri ES114 ([http://ijs.sgmjournals.org/content/57/12/2823 now Aliivibrio fischeri ES114]) and Streptomyces coelicolor A3(2) are environmental bacteria with well-characterized detection and catabolic cascades for chitin use as a carbon source. The way by which each bacteria detect the chitin is through a two-component system. Figure 1 illustrates the proposed sensing model for the system in A. fischeri, whose activation depends on the interaction CBP + N-acetyl-glucosamine starting the signaling from ChiS histidine kinase to turn on the chitinolytic genes. Figure 2 depicts how S. coelicolor uses a pretty similar system to the previously described, to phosphorylate ChiR regulator and induce transcription of the catabolism genes. In both cases, we expect to extract and put the genes into functional and independent biobricks available to further applications of chitin sensing and/or degradation.

Our bacteria

Aliivibrio fischeri ES114

Genes a usar: Diagramita del sistema:

Streptomyces coelicolor A3(2)

Genes a usar: Diagramita del sistema:

References

[1] [http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=312309&lvl=3&lin=f&keep=1&srchmode=1&unlock NCBI Taxonomy Browser: Aliivibrio fischeri ES114]

[2] [http://www.ncbi.nlm.nih.gov/Taxonomy/Browser/wwwtax.cgi?mode=Info&id=312309&lvl=3&lin=f&keep=1&srchmode=1&unlock NCBI Taxonomy Browser: Streptomyces coelicolor A3(2)]

[3] [http://www.pnas.org/content/101/2/627 Li, X., and Roseman, S. (2004). The chitinolytic cascade in Vibrios is regulated by chitin oligosaccharides and a two-component chitin catabolic sensor/kinase. PNAS 101, 627–631.]

[4] [http://mic.sgmjournals.org/content/154/2/373.full Colson, S., Wezel, G.P. van, Craig, M., Noens, E.E.E., Nothaft, H., Mommaas, A.M., Titgemeyer, F., Joris, B., and Rigali, S. (2008). The chitobiose-binding protein, DasA, acts as a link between chitin utilization and morphogenesis in Streptomyces coelicolor. Microbiology 154, 373–382.]