Weill Cornell Medicine investigators have revealed the detailed workings of a cell membrane protein that has essential roles in all animals. The discovery could lead to new therapeutic strategies for blood coagulation disorders, cancers and other conditions in which the protein, called a TMEM16 scramblase, works abnormally.
Scramblases operate within cell membranes, where they alter or "scramble" the normal layered arrangement of lipid molecules—an essential step in many biological processes. The scramblase TMEM16F also works as an ion channel, allowing small, charged molecules such as potassium or chloride ions through the membrane. Despite its broad importance, scientists have been unable to capture images of the functioning protein at high resolution. In the study, published April 17 in Nature Structural and Molecular Biology, the researchers at last attained this goal by embedding the protein in tiny lipid capsules called liposomes, which allowed them to image its active and inactive structures at near-atomic-scale resolution.
"We are excited by this finding because it paves the way for the targeted discovery of inhibitors or activators of this scramblase, which could be used, for example, to treat coagulation-related disorders," said study senior author Dr. Alessio Accardi , a professor of biochemistry and biophysics in anesthesiology at Weill Cornell Medicine.
The study's co-first authors were Accardi laboratory members Dr. Zhang Feng, a postdoctoral researcher, and Omar Alvarenga, a doctoral candidate in the Weill Cornell Graduate School of Medical Sciences .
TMEM16F's rearrangement of cell membrane lipids enables platelet cells to clump together to make blood coagulate, and a mutation affecting the scramblase underlies a hemophilia-like bleeding disorder called Scott Syndrome. The protein is also involved in the formation of the placenta in pregnancy, bone development and immune functions; and it is exploited or suppressed in various cancers and infections.
Cell biologists and drug developers have long wanted to apply high-resolution structural imaging techniques to learn precisely how TMEM16F works. But the scramblase's complex structure essentially falls apart when not embedded in a cell membrane, and scientists have had difficulty recreating that membrane environment when trying to image the protein in the laboratory.
In the study, Dr. Accardi's team overcame these difficulties by imaging copies of TMEM16F embedded in artificial liposomes—tiny, spheroid structures with layered lipid membranes very similar to those of a larger cell.
"The liposome membranes proved to be better mimics of the native cell-membrane environment of TMEM16F, and we were able to verify that the protein can be active in them," said Dr. Accardi, who is also a professor of biochemistry and biophysics at Weill Cornell Medicine.
High levels of calcium ions in the cell interior normally drive TMEM16F into its active, lipid-scrambling state. Dr. Accardi's team reproduced this condition in TMEM16F-containing liposomes and used low-temperature electron microscopy to compare normal and mutated versions of the protein. They found that elements of the protein rotate to form an X-shape in the membrane which opens a pore or groove. With help from computer modeling, they showed that ions can traverse the membrane moving through this pore and that this novel arrangement of the groove disrupts the normal lipid arrangements surrounding TMEM16F.
"Ions move through the inside of TMEM16F and lipids move on the outside," Dr. Accardi said.
He added that the revealed mechanism of TMEM16F activity is strikingly different from that seen in earlier studies of related scramblases.
Having such detailed "pictures" of TMEM16F's key conformations means that researchers for the first time can design drug molecules specifically to target them.
"An activator of TMEM16F could be used, for example, as a pro-coagulant to treat patients with Scott Syndrome or other bleeding disorders, while an inhibitor could be used as an anticoagulant—among many other potential applications," Dr. Accardi said.
The research reported in this story was supported by in part by the National Institute of General Medical Sciences, part of the National Institutes of Health, through grant R35 GM152012.
