A protein that is a key modulator of fat, glucose and cholesterol levels in the body usually works in tandem with another protein, but new research shows it can also work with an unexpected partner - itself. A team of Penn State researchers has now characterized the structure of this twin pairing, finding that while its conformation is different, it can still perform its function of activating the expression of other genes. The finding, the team said, could open new pathways for therapies for liver cancer, diabetes and other metabolic diseases by targeting the twin pair to treat or prevent disease with potentially fewer off-target side effects.
A paper describing the research published this week (Feb. 23) in the journal Nucleic Acids Research.
The farnesoid X receptor (FXR) protein is found primarily in the liver, kidneys and intestine, where it helps maintain the balance of lipid, glucose and bile acid levels. It predominantly functions by partnering with another protein called the retinoid X receptor alpha (RXR). Together, the two proteins bind to specific sequences of DNA and function as a receptor for molecules known as ligands. Once bound by ligands, the complex of molecules can recruit cellular machinery to turn on or off genes involved in synthesizing bile acids to regulate lipid and glucose metabolism.
"FXR has been implicated in metabolic disease and in some gastrointestinal and liver cancers," said research team leader Denise Okafor, Dorothy Foehr Huck and J. Lloyd Huck Early Career Chair in Biophysics, assistant professor of biochemistry and molecular biology, and of chemistry in the Penn State Eberly College of Science. "Targeting the FXR-RXR complex as a therapeutic can be problematic because RXR, like a Swiss army knife, performs many different functions by partnering with other receptor proteins. So, if its function is disrupted it could lead to unintended off-target consequences. Previous studies have hinted that FXR can also partner with another molecule of FXR, so we were interested in what the structure of this FXR-FXR complex looked like and if it could still function to drive gene expression and possibly be a new target for the design of therapeutics."
In the lab, the researchers combined purified FXR with segments of synthetic DNA that included the specific sequences to which FXR normally binds. They confirmed previous reports that FXR can bind to DNA as an individual molecule or as a pair. In a separate experiment, the team then demonstrated that the FXR-FXR pairing could recruit and bind to the required cellular components and drive gene expression.
"We then used an imaging technique called small-angle X-ray scattering to characterize the three-dimensional structure of the FXR-FXR complex," said Sabab Hasan Khan, assistant research professor of biochemistry and molecular biology and the first author of the paper. "We found that the molecules take a very different conformation in comparison to the FXR-RXR complex. The molecules become extended and, unlike any previously characterized receptor protein pairings, the regions of the proteins that bind to ligands are separated and do not interact with each other like they do in the FXR-RXR pair."
This unusual conformation suggests that the FXR-FXR pairing may functionally drive the expression of a different set of genes than the usual FXR-RXR pair, according to the researchers. Further research could explore the which genes and the potential separate roles of the two types of pairings.
"We could be uncovering a hidden function of this receptor that has been masked all these years because we thought its function was defined by its partnership with RXR," Okafor said. "Because FXR is so fundamentally involved in the liver and liver disease, diabetes and other metabolic diseases, there's just so much to understand about this newly characterized structural variant. What genes does it regulate? Are these genes involved in different pathways? Are there different cellular processes that are being regulated or modulated? Answering these questions could tell us that there's a lot of other biology that hasn't yet been explored. But it also opens the door for new approaches to treating or preventing diseases associated with FXR."
In addition to Okafor and Khan, the research team at Penn State included Neela Yennawar, director of the Biomolecular Interactions Core Facility, co-director of the X-Ray Crystallography and Scattering Core Facility and research professor in the Penn State Huck Institutes of the Life Sciences.
The U.S. National Institutes of Health, the U.S. National Science Foundation and the Penn State Huck Institutes of the Life Science funded the research.
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