Cholesterol plays a central role in cell health, influencing membrane fluidity, curvature, and signaling. To maintain proper cholesterol levels, cells rely on tightly regulated transport systems that move cholesterol between membranes and organelles. However, many of the proteins involved in sensing, binding, and trafficking cholesterol—and how they dynamically interact—remain poorly understood.
A team of researchers, led by Professor Keriann Backus and chemistry and organic chemistry Ph.D. candidate Andrew P. Becker, has developed a powerful new proximity labeling tool that reveals how cholesterol and proteins interact inside living cells—offering fresh insights into the molecular machinery that governs cellular cholesterol balance and membrane biology.
The study, published in a recent issue of Nature Chemical Biology, introduces POCA (photosensitizer-dependent oxidation and capture by amine), a singlet oxygen–based proximity labeling platform that can map both protein and lipid interactomes inside cells. Unlike existing methods, which typically rely on separate strategies to study protein–protein and protein–lipid interactions, POCA enables the identification of both types of interactions in their native cellular context.
POCA addresses existing gaps in proximity labeling by using cell-penetrant photosensitizer reagents that can be easily incorporated to small-molecules—such as cholesterol—or proteins via the widely used HaloTag system. When activated by light, photosensitizers generate singlet oxygen,a reactive species that labels nearby proteins, allowing researchers to identify the proteins interacting with the tagged cholesterol or HaloTag proteins within the cell.
Using cholesterol-directed POCA, the researchers successfully captured both known and previously unrecognized cholesterol-binding proteins. Notably, the platform identified protein complexes whose interactions changed in response to intracellular cholesterol levels, as well as proteins uniquely associated with cholesterol delivered through natural pathways such as lipoprotein uptake—a key advance over existing cholesterol probes that rely on artificial delivery methods and have much lower signal. Beyond individual proteins, POCA exposed cholesterol-sensitive behavior in larger assemblies, including the ER membrane complex (EMC), identifying it as a cholesterol-responsive protein complex for the first time.
The team also paired POCA with the widely used HaloTag system, demonstrating the method’s adaptability for protein-directed interactomics. Applying this approach to Aster-B, a cholesterol transport protein that moves cholesterol from the plasma membrane to the endoplasmic reticulum (ER), the researchers uncovered sterol-dependent changes in Aster-B’s interaction network. They further revealed domain-specific crosslinking within Aster proteins and unexpected localization of Aster complexes within flotillin-rich detergent-resistant membranes, suggesting new layers of regulation in cholesterol transport.
Overall, the study establishes POCA as a straightforward, broadly applicable interactomics platform that overcomes long-standing limitations in proximity labeling technologies. By enabling unified, in situ mapping of protein and lipid interactions using readily available tools, POCA opens new avenues for studying membrane biology, cholesterol homeostasis, and dynamic cellular interactions relevant to human health and disease.
The study represents a collaborative effort across multiple UCLA departments and institutes as well as the University of Tokyo. The paper’s first author is Chemistry & Biochemistry Ph.D. student Andrew P. Becker, and co-authors are Elijah Biletch, John Paul Kennelly, Soon-Gook Hong, Ashley R. Julio, Miranda Villanueva, Rohith T. Nagari, Daniel W. Turner, Nikolas R. Burton, Tomoyuki Fukuta, Liujuan Cui, Xu Xiao, Zaid Vellani, Alexander Nguyen, Julia J. Mack, Peter Tontonoz, and senior author Keriann M. Backus.
Backus joined the UCLA faculty in 2018 as an Assistant Professor of Biological Chemistry in the UCLA David Geffen School of Medicine, with a joint appointment in the UCLA Department of Chemistry & Biochemistry. In 2020, she was appointed to UCLA’s Alexander and Renee Kolin Endowed Professorship of Molecular Biology and Biophysics. She was promoted to Associate Professor with tenure in July 2024.
Backus’ innovative research has earned her numerous awards and recognitions, including the Packard Fellowship, NIH New Innovator Award, Beckman Young Investigator Award, Ono Pharma Foundation Breakthrough Science Initiative Award, DARPA Young Faculty Award, and the International Chemical Biology Society (ICBS) Young Chemical Biologist Award.
Prior to UCLA, Becker worked as a process chemist at the pharmaceutical company Neurocrine Biosciences where he developed the manufacturing process for the small molecule Crinecerfont, which received FDA approval in December 2024. He began his Ph.D. in 2020 at UCLA with the goal of applying synthesis to solve problems in biological systems. He has been supported by the Stone and Senior Foote Fellowships at UCLA.
Penny Jennings, UCLA Department of Chemistry & Biochemistry, penjen@g.ucla.edu.
