Molecular and cellular biology graduate students Ben Johnson and Mariana Breña, with their advisor, professor of biochemistry and molecular biology Alejandro Heuck, recently published a paper in the Springer Nature journal Scientific Reports in which they describe how they developed a new molecular probe for measuring the amount of accessible cholesterol in cells. The work was part of Johnson’s Ph.D. dissertation; he is now a postdoctoral researcher at the U.S. Army’s Natick Soldier Research, Development and Engineering Center.
While most people think of cholesterol in narrow terms as a molecule linked to heart attack risk, it is in fact found in every cell of the body. Heuck explains, “Cholesterol is essential to life, we cannot live without it, and diseases such as atherosclerosis arise when cells lose control of the proper balance of cholesterol levels.”
He and colleagues have for many years studied how toxic proteins secreted by pathogens make holes in cell membranes, in particular a toxin secreted by the gas gangrene causative pathogenClostridium perfringens. This toxin, Perfringolysin O, targets cholesterol in the cell membrane to form its holes. Heuck’s fundamental work on this process led him to realize that because the toxin recognizes cholesterol as a binding target, Perfringolysin O can serve as a cholesterol biosensor. Using an inactivated version of Perfringolysin O, he, Johnson and Breña developed just that.
As Heuck explains, the cholesterol research community once focused only on the total amount of cholesterol present in the membrane, and how this amount affected cholesterol transport in the cell. “People measured total cholesterol in order to study how it interacts with other molecules and how it gets in and out of the cell,” he says. But it is becoming clear that knowing total cholesterol is not enough, he adds. “We needed to determine how much of the total cholesterol is accessible for transport. We are starting to realize that accessibility of cholesterol plays an important role in how cholesterol levels are regulated in the cell.”
Heuck points out, “With the tools previouly available to measure total cholesterol, it was very difficult to measure changes in cholesterol accessibility. The biosensor we’ve develpoed allows us to detect changes in cholesterol accessibility and to generate fine-tuned probes, resulting in a palette of biosensors to measure slight changes in cholesterol levels. We have improved the probe for the study of live cells and defined the proper way to use the probe to measure changes in cholesterol accessibility. It will be useful for anyone in the field of cholesterol transport and cholesterol-related diseases, which is a large area of research. I expect that scientists will take advantage of the new biosensors and what we have learned about them.”