Scientists have discovered how a toxin produced by a common intestinal bacteria gains access to colon cells, solving a mystery that has baffled researchers for more than 15 years. The discovery not only explains how the toxin begins to damage the colon, but also points to a possible new way to block its effects before they contribute to colorectal cancer.
The findings come from a multi-institutional team led by researchers at the Johns Hopkins Kimmel Cancer Center, Bloomberg ~ Kimmel Institute for Cancer Immunotherapy, and the Johns Hopkins University School of Medicine. Published in NatureThe study shows that the toxin, known as BFT and produced by Bacteroides fragilisIt must first bind to a host protein called claudin-4 before it can damage colon cells. The research was supported in part by the National Institutes of Health.
“We’ve made several attempts over time to identify the receptor, so this is an exciting time,” says senior author Cynthia Sears, MD, Bloomberg ~ Kimmel Professor of Cancer Immunotherapy and professor of medicine at Johns Hopkins. “Understanding how bacterial toxins work may open the door to new approaches to detecting and treating associated diseases, such as diarrhea, colorectal cancer, and bloodstream infections.”
A hidden receptor gives intestinal toxins access to colon cells
The team’s findings have already inspired a promising strategy to block the toxin. The researchers developed a molecular decoy that successfully intercepted BFT in animal models, preventing it from damaging the colon.
Bacteroides fragilis It is found in up to 20% of healthy people, but certain strains can cause inflammation in the colon and promote tumor growth. Previous research from the Sears lab showed that BFT causes chronic inflammation by cutting off E-cadherin, a protein that helps maintain the colon’s protective barrier. that before Nature medicine The study also showed that the toxin’s activity drives the formation of colon tumors.
An important question remained unanswered. BFT did not appear to bind directly to E-cadherin, suggesting that another molecule first helped the toxin access its target.
CRISPR screen reveals missing link
To identify that missing piece, Maxwell White, MD/Ph.D. candidate in the Sears lab, led a genome-wide CRISPR screening effort in collaboration with Matthew Waldor’s lab at Harvard Medical School.
The researchers systematically turned off individual genes in colonic epithelial cells to determine which ones were necessary for the toxin to work. One protein stood out immediately: claudin-4. When claudin-4 was removed, BFT could no longer adhere to cells, leaving E-cadherin unharmed.
“It took us a while to get the assay working and validate the approach, but once we were able to run the analysis, Claudin-4 was a clear and resounding success,” White says. “That was an exciting moment.”
The discovery surprised the researchers. Sears says many scientists expected the receptor to be a signaling protein, like a G-coupled protein receptor, but claudin-4 belongs to a different class of proteins. A review of previous research also failed to discover another toxin that behaves in the same way. Most protease toxins bind directly to the molecules they attack rather than first binding to a separate receptor.
Scientists confirm the molecular target of the toxin
To verify the interaction, the Johns Hopkins researchers teamed up with structural biologists F. Xavier Gomis-Rüth and Ulrich Eckhard from the Barcelona Institute of Molecular Biology.
Using biophysical techniques, White and the Barcelona team showed that BFT and claudin-4 form a tightly bound one-to-one complex in laboratory experiments. This provided the first direct physical evidence that the toxin binds to the receptor before damaging colon cells.
The researchers then tested their findings in living systems through a collaboration with Min Dong’s lab at Harvard Medical School. Working with Kang Wang and his colleagues, they examined how the toxin behaved in mouse models.
Molecular decoy protects mice from intestinal toxin
The team created a soluble version of claudin-4 that acted as a decoy by displaying portions of the receptor normally recognized by the toxin. Instead of binding to colon cells, BFT bound to decoy proteins.
This strategy successfully protected mice from BFT-induced colon damage.
“This approach could be repeated with small molecules or other biologics that have better pharmacological properties,” says White. The team is now investigating what types of therapies may be most effective in blocking the toxin.
There are still questions
Although the researchers identified the receptor and showed that it binds closely to BFT, a major challenge remains unsolved. They have not yet captured the precise experimental structure that shows exactly how the toxin and claudin-4 fit together.
Current AI modeling tools, including AlphaFold, were unable to fully resolve the interaction.
Other authors of the paper include Jason Chen, Shaoguang Wu, Abby L. Geis and Jessica Queen of Johns Hopkins and Hailong Zhang, Karthik Hullahalli and Jie Zhang of Harvard Medical School.
The research was supported by the Bloomberg~Kimmel Institute for Cancer Immunotherapy, Janssen Research and Development, Cancer Research UK, the National Institutes of Health (grant numbers R01 AI042347, R01 NS080833, R01 NS117626, R01 AI170835, and R01 AI189789), and the Howard Hughes Medical Institute.
Sears receives royalties for writing and reviewing for UpToDate. This agreement is managed by Johns Hopkins University in accordance with its conflict of interest policies.