Using a combination of new technologies and state-of-the-art protein sequencing techniques, a research team led by scientists from the Whitehead Institute for Biomedical Research has painted a new picture of the human cell membrane.
Their work, described in the cover story of the July 1994 issue of The Journal of Cell Biology, has specific implications for the prevention of atherosclerosis, complications of diabetes, and some common effects of aging. The findings also suggest new approaches to therapy for diseases ranging from cancer to cystic fibrosis.
"Every interaction between a cell and its environment must take place through the cell membrane," said Dr. Michael Lisanti, a Whitehead Institute Fellow and the first author of the paper. "We've known for years that specific proteins in the membrane pick up signals from the environment, and that other proteins transmit those signals across the membrane into the cell, but we didn't understand how the whole system fit together. We tended to envision a chaotic scene much like a bumper car ride, in which the molecule responsible for capturing a signal raced around the surface of the cell until it bumped into the correct transmitter protein and then proceeded to hand over the message.
"In retrospect, we should have known that the system could not be that disorganized," he added.
Dr. Lisanti and his colleagues first began to suspect a new order in the world of membrane biology last year, when they isolated a class of cell organelles called caveolae, or "little caves." Now they have identified the components of the caveolae and discovered a host of familiar protein molecules bound together in a compact depot or message center optimally designed to process signals from the outside world.
"When a receptor molecule captures a specific signal from the environment, it doesn't have to search all over the surface of the cell for the appropriate transmitter protein," Dr. Lisanti said. "It simply delivers the signal to a nearby caveola, where a broad range of transmitters are ready and waiting."
Among the proteins found in the caveolae are membrane-spanning proteins implicated in the development of atherosclerosis, diabetic vascular complications, normal aging and cancer; gateway proteins that regulate the passage of small molecules into the cell (manipulation of these gateway molecules in caveolae could provide new avenues for drug delivery, especially chemotherapy), and a variety of signaling proteins involved in normal cell growth and cancer. Thus, the isolation of caveolae offers a wealth of new targets for the prevention, diagnosis and treatment of human disease.
Atherosclerosis, or hardening of the arteries, occurs when cholesterol-bearing molecules build up just below the layer of specialized cells lining the inner surface of blood vessels. Many theories have been proposed to explain why this happens, including the idea that the lining cells must be damaged in some way to allow the build-up to occur.
"The problem has been that physicians often see atherosclerosis when there is no detectable damage to the cells," Dr. Lisanti said. "Some other process must be at work. Our analysis of the contents of caveolae has revealed high concentrations of a molecule called CD 36, which is known to bind oxidized LDL (Low-Density Lipoprotein), the cholesterol-bearing molecule most often implicated in atherosclerosis."
Dr. Lisanti and his associates suspect that CD 36 inside the caveolae binds oxidized LDL from the bloodstream and prepares it for transport. The oxidized LDL is transcytosed; that is, it is packaged in little compartments, carried through the specialized cells lining the blood vessel, and then deposited on the other side. The isolation of caveolae will greatly facilitate the search for new ways of blocking this process, thereby preventing atherosclerotic build-up.
Another major component of the caveolae is a molecule called RAGE (receptor for advanced glycosylation end products, or AGE's). RAGE binds molecules in the bloodstream that have been altered or damaged by prolonged exposure to blood glucose. Abnormal accumulation of such molecules has been implicated in diabetic vascular complications and in normal aging.
"Again, we believe that caveolae may play a central role in this process," Dr. Lisanti said. He explained that RAGE in the caveolae of endothelial cells lining the small blood vessels may concentrate the glucose-altered molecules, resulting in abnormal deposits inside the blood vessel walls. Further research on caveolae may reveal a way to block this process to prevent the thickening of blood vessel walls that leads to destruction of the retina, kidneys and other organs in diabetics.
Gateway Proteins and Cystic Fibrosis
The caveolae also contain a third transmembrane molecule with an ancient history. This is a "window" on the cell, a tiny portal that may allow the passage of rare nutrients and other molecules through the cell membrane. The portal, called "porin," closely resembles similar structures found in bacteria, suggesting that the caveolae may be very old structures in the evolution of life, Dr. Lisanti noted.
The discovery of porin in caveolae has many different implications for medical science. First, there is evidence from other investigators that porin interacts with the cystic fibrosis protein (CFTR), making it an ideal target for further research on the origins of cystic fibrosis. Second, porin may be responsible for the uptake of folate, a vitamin required for DNA synthesis. This is important because one of the most common anti-cancer drugs is the anti-folate compound methotrexate. Studies of porin function in caveolae may help explain why some tumors become methotrexate-resistant and suggest new strategies for overcoming this resistance.
Finally, the porin molecule makes a relatively large hole in the cell membrane inside the caveolae. If scientists could learn how to open and close these portals at will, they would have an important new avenue for introducing therapeutic drugs. This work could lead, in turn, to new strategies for crossing the blood-brain barrier, the physiological boundary that often blocks drug delivery to the central nervous system. (The blood-brain barrier consists primarily of endothelial cells which, in other tissues, are known to be rich in caveolae.)
The analysis of caveolar proteins also indicates that the caveolae play a central role in normal and abnormal cell growth. In mammalian cells, growth must be regulated very precisely to maintain proper development of organs and tissues. Cells send and receive a constant barrage of "growth" and "no growth" signals in the form of hormones and other chemical messengers to ensure that they are acting in concert with other cells in the body. When this message system breaks down (because of some genetic accident or an adverse environmental effect), the result is unregulated growth, or cancer.
In their new paper, Dr. Lisanti and his colleagues report evidence that the caveolae play a vital role as switchboards for incoming growth-related signals. The caveolae contain major elements of the complex network required to transmit external growth-related signals to the cell's internal regulatory machinery, including known tumor suppressor proteins and potential oncogenes.
"This finding could help identify new targets for cancer therapy," Dr. Lisanti said. "It should simplify the task of deciphering how different parts of the network interact, and suggest new ways for blocking that interaction in tumor cells."
The authors of the JCB paper in addition to Dr. Lisanti were Philipp E. Scherer, ZhaoLan Tang, Ya-Huei Tu, and Massimo Sargiacomo of the Whitehead Institute; Richard F. Cook, Director of the Biopolymers Laboratory, Howard Hughes Medical Institute, Center for Cancer Research, Department of Biology, MIT; Jolanta Vidugiriene of the Laboratory of Molecular Parasitology, The Rockefeller University; and Anne Hermanowski-Vosatka of Brigham and Women's Hospital and Harvard Medical School.
This work was supported by a National Institutes of Health (NIH) FIRST Award to Dr. Lisanti; a grant from the W.M. Keck Foundation to the Whitehead Fellows Program; a fellowship from the European Molecular Biology Organization to Dr. Scherer, and additional NIH grants to the Whitehead Institute.
A version of this article appeared in the July 20, 1994 issue of MIT Tech Talk (Volume 39, Number 1).