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Bacterial-Epithelial Interactions in the Mammalian Gut
Summary: Lora Hooper is investigating how the intestinal immune system defends against the vast microbial communities that inhabit the mammalian gut.
Humans harbor nearly 100 trillion intestinal bacteria that are essential for health. Millions of years of coevolution have molded this human-microbe interaction into a symbiotic relationship. Intestinal bacteria benefit their human hosts by increasing digestive efficiency and helping to extract key nutrients from the ingested diet. In return, these microbes are given safe haven in a nutrient-rich environment.
Epithelial cells that line the intestinal surface are in constant contact with these vast microbial communities, and thus constitute the major interface between the microbiota and internal host tissues. Although intestinal microbes perform beneficial functions, they can still cause disease if they cross intestinal epithelial barriers and invade deeper host tissue. However, microbial incursions across the intestinal surface are relatively rare, despite the enormous numbers of bacteria present in the gut. Thus, the intestinal epithelium likely plays a key role in sequestering bacteria in the gut lumen and preventing their penetration of host barriers.
A major goal of my laboratory is to understand how gut epithelial cells "keep the peace" with resident microbes. To deepen our understanding of these interactions, we use a multidisciplinary approach that includes biochemical analyses of new antibacterial proteins, in vivo analyses of the pathways by which gut epithelia sense bacteria, and creation of animal models to determine how epithelia regulate microbial interactions with host tissues. We hope that our studies will lead to fundamental insights into bacterial-epithelial interactions and will affect our basic understanding of how mammals maintain symbiotic relationships with their indigenous microbial communities.
Germ-Free Mice as a Tool for Analyzing Intestinal Host-Microbial Relationships
Understanding the molecular foundations of intestinal host-microbial relationships is a challenging problem due to the complexity of the gut microflora, the complexity of the intestinal mucosal surface, and the lack of good in vitro models for studying the complex interplay between host and microbe. Overcoming these challenges has required the development of unique in vivo experimental approaches. One of our most important tools is gnotobiotics ("known life"), a technology involving the use of microbiologically sterile (germ-free) animals. Using germ-free mice, we can manipulate the intestinal ecosystem by introducing a single bacterial species or defined species mixtures. In my laboratory, we routinely combine gnotobiotics with a battery of tools for molecular analysis to study bacterial-epithelial crosstalk in the gut. Thus far we have discovered a new mechanism of innate mucosal defense and essential insights into how gut epithelia regulate interactions with intestinal bacteria in vivo.
Epithelial Antimicrobial Proteins
Much of our work has focused on secreted epithelia-derived antimicrobial proteins, which directly lyse bacteria and likely promote symbiotic host-microbial relationships by restricting bacterial contact with host tissues. In my early studies of intestinal host-microbial interactions, I discovered that colonization of germ-free mice with commensal microbes triggers epithelial expression of angiogenin-4 (Ang4), an RNase that is targeted to the gut lumen and directly lyses intestinal bacteria. This work revealed a novel mechanism of innate mucosal defense, and showed for the first time that expression of epithelial bactericidal proteins can be elicited by gut bacteria. Subsequently, we have used gnotobiotics, laser capture microdissection, and DNA microarrays to search for new members of the microbe-inducible antibacterial arsenal of gut epithelia. These efforts led to the discovery of RegIIIγ, a secreted member of the C-type lectin family of carbohydrate-binding proteins. We found that RegIIIγ is discharged into the gut lumen and mediates direct killing of Gram-positive gut bacteria. As RegIIIγ is the first example of a directly bactericidal lectin, it represents a previously unappreciated biological function for the C-type lectin family.
We gained insight into the biochemical basis of RegIIIγ bactericidal activity by showing that its ligand is peptidoglycan, a major component of Gram-positive bacterial surfaces. We found that RegIIIγ binds peptidoglycan through interactions with the glycan moiety, suggesting a novel mechanism of peptidoglycan binding. RegIIIγ is distinct from other C-type lectins in that it is specific for polysaccharide ligands and will not bind monosaccharides. This is likely an important factor in allowing RegIIIγ to distinguish between bacterial and mammalian cell surfaces, since the extended glycan structures of peptidoglycan are unique to microbial cell walls. We are using biochemical and structural approaches to understand the RegIIIγ-peptidoglycan interaction and thus gain molecular insight into how RegIIIγ targets gut bacteria for killing.
Epithelial Contributions to Intestinal Host-Microbial Homeostasis
Although the intestinal epithelium is in direct contact with the vast microbiota, little is known about how epithelial cells defend the host against the heavy bacterial load. We are studying this question by focusing on the Paneth cell. The Paneth cell is a specialized small intestinal epithelial lineage that resides at the base of crypts of Lieberkühn and contributes to intestinal innate immunity through the secretion of antimicrobial proteins. We are using powerful tools for the genetic manipulation of this epithelial lineage to uncover the mechanisms by which Paneth cells sense intestinal bacteria. In addition, we are using an in vivo lineage ablation model to determine how this specialized epithelial cell functions to maintain intestinal host-microbial homeostasis. We anticipate that this work will yield general insights into how intestinal epithelial cells regulate host-bacterial interactions at the mucosal interface.
This work is also supported by the National Institutes of Health, the Crohn's and Colitis Foundation of America, the Burroughs Wellcome Fund, and the Welch Foundation.
Last updated August 12, 2008
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