Biosynthesis of Nucleotide Sugar Monomers for Exopolysaccharide Production in Myxococcus Xanthus
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Myxococcus xanthus displays social (S) motility, a form of surface motility that is key to the multicellular behaviors of this organism. S motility requires two cellular structures: type IV pili (TFP) and exopolysaccharides (EPS). Previous studies have shown that M. xanthus does not use glucose or any other sugar as a primary carbon source. However, eight monosaccharides, namely glucose, mannose, arabinose, galactose, xylose, rhamnose, N-acetyl-glucosamine, and N-acetyl-mannosamine, are found in M. xanthus EPS. In this study, pathways that M. xanthus could use to produce the activated sugar monomers to form EPS are proposed based on genomic data. Of the eight sugars, pathways for seven were disrupted by mutation and their effects on the EPS-dependent behaviors were analyzed. The results indicate that disruption of the two pathways leading to the production of activated rhamnose (GDP- and TDP-rhamnose) affected fruiting body formation (GDP form only) and dye binding ability (both forms) but not S motility. Disruptions of the xylose, mannose, and glucose pathways caused M. xanthus to lose S motility, fruiting body formation, and dye binding abilities. An interruption in the pathway for galactose production created a mutant with properties similar to a lipopolysaccharide (LPS) deficient strain. This discovery led us to study the phenotypes of all mutant strains for LPS production. The results suggest that all mutants may synthesize defective LPS configurations. Disruption of the UDP-N-acetyl-mannosamine pathway resulted in a wild type phenotype.
In addition, it was discovered that interruption of the pathway for N-acetyl-glucosamine production was possible only by supplementing this amino-sugar in the growth medium. In an attempt to determine if other mutants could be recovered by sugar supplementation, it was discovered that the Î pgi mutant can be rescued by glucose supplementation. The Dif chemotaxis-like pathway is known to regulate EPS production in M. xanthus. DifA is the upstream sensor of the pathway. Previous studies had created a NarX-DifA chimeric protein, NafA, that enables the activation of the Dif pathway by nitrate, the signal for NarX. In this study, we constructed a Î pgi difA double mutant containing NafA. This strain was then subjected to various incubations with glucose and/or nitrate to determine whether the point of EPS regulation by the Dif pathway is down- or up-stream of the step catalyzed by Pgi (phosphoglucose isomerase). Preliminary results from this study are inconclusive.
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