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A Key Control Knob for Microbiome Metabolism

Researchers are uncovering the complex mechanisms by which human gut bacteria accomplish a variety of functions, from transforming compounds derived from food into nutrition for the host to producing molecules that impact human behavior and performance.

The Venturelli Lab has now overturned prior knowledge about how polysaccharide utilization loci (PULs) mediate the fitness and community-level interactions of Bacteroides uniformis, one of the most abundant species of bacteria in the human gut. The researchers detail their groundbreaking methods and results in the February 2022 issue of Cell Host & Microbe.

Bacteroides uniformis relies on clusters of genes called PULs to determine its ability to persist in the gut (also known as fitness) and modulate community-level interactions between other microbes in the gut. PULs enable Bacteroides to produce a suite of enzymes that degrade diet and host-derived glycans — carbohydrate-based polymers made by all living organisms that are also known as polysaccharides — into metabolites that impact the host metabolism and physiology as well as the growth and metabolic activities of the gut ecosystem.

“We get around 10% of our energy from metabolites produced by gut microorganisms, and polysaccharide utilization loci are major determinants of these chemical transformations. But we still don’t fully understand the function of the majority of PULs and how PULs influence microbial interactions in the gut,” says Jun Feng, the lead author on the Cell Host & Microbe study. Feng is an assistant scientist working in the Venturelli Lab.

To improve our understanding of how PULs influence the fitness of bacteria and interactions with other microbes and their host, Feng and his colleagues studied 23 of the 55 PULs in Bacteroides uniformis in the presence of different nutrients in vitro and in vivo. Their comprehensive study on the fitness of Bacteroides uniformis provides a deeper understanding of how diverse PULs contribute to glycan utilization, microbial interactions, and bacterial colonization of the mammalian gut.

“The approach we took was to really focus and examine the effect of complex glycan degradation pathways, which are critical to the gut microbiome,” says senior author and biochemistry assistant professor Ophelia Venturelli.

By introducing different PULs deletion mutants (PULs that are missing metabolic pathways) into a microbial community created in the lab, the researchers found that PULs can benefit or harm bacterial colonization depending on specific conditions in the microbial community. PULs also may influence global gene expression patterns in the gut. The researchers designed a barcoding approach to tag the PULs before they were introduced to the synthetic microbial community, which allowed them to carefully study these interactions and impacts.

“That’s the most important finding of this study,” said Feng. “Previously, people thought that polysaccharide utilization loci always promoted bacterial colonization. We found that’s not true: there can be a fitness benefit or cost depending on the environmental condition.”

The researchers also studied diet-dependent microbial interactions in vivo by feeding germ-free mice one of three controlled diets. The diets — a higher-fiber diet, a low-fiber diet, and a high-fat diet — varied in the type and abundance of carbohydrates accessible to bacteria in the gut. Feng and his colleagues found that the abundance of Bacteroides uniformis in the mouse gut was highest in mice fed a high-fiber diet and significantly lower in mice receiving a low-fiber diet. The high-fat diet group also exhibited differences in gut microbiota compared to the low-fiber diet group, and temporal changes in the composition of the microbial community varied across diets.

Results of these in vivo experiments suggest that PULs play a major role in the ability of Bacteroides uniformis to colonize and persist in the gut: diet is yet another variable that may influence whether PULs provide a benefit or cost to microbial fitness.

Finally, to better understand how PULs modulate the growth of butyrate-producing bacteria (butyrate is produced by specialized gut bacteria that promote gut health as an end product of fermentation), Feng, in close collaboration with another assistant scientist in the Venturelli Lab, modeling expert Yili Qian, studied combinations of microbe-microbe interactions with specialized butyrate producing bacteria and the downstream effects of these interactions. Their combined experimental and computational results, the researchers say, suggest different classes of mechanisms by which PULs and metabolic byproducts of Bacteroides uniformis can influence interspecies interactions and butyrate production.

“The results of this study are very interesting and provide more of a systems-level understanding of how polysaccharide utilization loci are shaping critical, health-relevant functions in the gut,” says Venturelli. “But the tools that Jun developed are also very useful for the scientific community.”

For example, the team’s Cell Host & Microbe study presents the first published application of CRISPR to genomic editing of Bacteroides.

“These tools are going to accelerate and enable genetic manipulation of important health-relevant bacterial species for a variety of health applications,” Venturelli says. “They allowed us to examine quantitatively how these pathways influence microbial communities in a variety of different contexts.”

Feng agrees. “I like to emphasize that Bacteroides uniformis is the most abundant bacteria in the gut, so a study on this is very important. We systematically developed genetic tools, including CRISPR-based genomic editing tools, and we also used a barcoding approach to tag each PUL mutant and study the behavior of each mutant in the community,” he says. “We found that the effects of polysaccharide utilization loci on Bacteroides colonization are complex. We did a lot in this paper, and this work and these tools will make a huge impact in the field.”

The study has also unlocked avenues for future investigation. The Venturelli Lab is currently investigating how specific PUL mutants contribute to high Bacteroides uniformis colonization ability. They’re interested in understanding how PULs are connected to one another and what causes the global changes in gene expression patterns that the researchers observed. And they’re also looking at interactions in larger communities that are more representative of what’s happening in the human gut.

“Once we understand these mechanisms at the genetic and molecular levels, we can start to devise strategies to manipulate gut ecosystem functions using tools from synthetic biology, protein engineering or chemistry,” Venturelli says. “Thinking about PULs as a key control knob for microbiome metabolism by modulating the production and degradation of health-relevant metabolites in the gut for benefiting human health holds tremendous promise for personalized medicine.”

Written by Catherine Steffel, Ph.D.

Graphical abstract describing three parts of the research study
Graphical abstract describing the Venturelli Lab's study.