Mechanistic studies to develop a polysaccharide degradation signature (PDS) and its application in improving host health

PROJECT SUMMARY/ABSTRACT
The human colon harbors a large number of microorganisms that collectively are referred to as the colonic
microbiome. The microbes in the colonic microbiome are dominated by bacteria of the phyla Bacteroidetes and
Firmicutes. Among the Bacteriodetes, Prevotella spp. and Bacteroides spp. abound in the colonic environment
and have evolved a complex protein machinery that allows them to harvest energy from both host undegradable
polysaccharides in the diet and host derived-glycans, such as mucin. Central to the mechanism underlying
polysaccharide degradation by the Bacteroidetes is the Polysaccharide Degradation Locus (PUL) or Loci (PULs)
present on their genomes. The PULs are composed of gene clusters that encode proteins that enable the
Bacteroidetes to sense, transport, and degrade diverse polysaccharides to their unit sugars for fermentation. A
large protein, known as the Hybrid Two Component System (HTCS), is conserved in the PULs of the
Bacteroidetes and functions by sensing either a polysaccharide or its oligosaccharides to turn on the expression
of the hydrolytic enzymes and their associated transporters. In this proposal, we demonstrate that indeed the
Bacteroidetes HTCS contain sensor modules that sense unique polysaccharides or their degradative products
in the colonic environment. Thus, we hypothesized that the diverse sensors in the HTCS polypeptides collectively
can serve as a proxy for polysaccharide sensing in the colon of an individual. We have designated this proxy as
the Polysaccharide Degradation Signature or PDS. By using more than 3000 HTCS sequences in the publicly
available databases, we constructed a phylogenetic tree that appeared to cluster the sensor modules into
different branches. Among host undegradable polysaccharides found in human diets, such as wheat, barley, rice
and oats, is arabinoxylan. We, therefore, used growth on arabinoxylan and transcriptomic analysis to determine
the PULs that target soluble arabinoxylan and insoluble arabinoxylan degradation, respectively, in three
members of the human colonic Bacteroidetes. Our data showed that clusters in our phylogenetic tree or PDS
can be matched to arabinoxylan sensing and metabolism. Interestingly, we also discovered that the Bacteroides
spp that metabolize complex arabinoxylan release the plant phenolic compound ferulic acid and that the
compound accumulates in the spent medium. Ferulic acid is known to have antioxidant effects and also to protect
against mechanosensory hair loss. We will, therefore, determine whether a synbiotic of complex arabinoxylan
and arabinoxylan-metabolizing Bacteroidetes has the capacity to confer protection against mechanosensory hair
loss in germ-free zebrafish. Confirmation of this observation will allow us, through transcriptomics analysis, to
determine the underlying molecular mechanisms for this protection. Furthermore, we will use biochemical and
structural analyses to completely characterize the mechanism of arabinoxylan degradation by the human colonic
Bacteroidetes. We also anticipate that our development of the PDS will allow rational manipulation of the
polysaccharides sensed by an individual’s microbiome for health and nutritional benefits.

Grant Number: 5R01GM140306-04
NIH Institute/Center: NIH
Principal Investigator: ISAAC CANN