March 10, 2005 — Scientists may have found a way to throw a wrench in the transmissions of several speed demons of the parasite world.
Researchers at Washington University School of Medicine in St. Louis and Harvard University have identified a protein that could help them develop drugs to stop or slow cell invasion by malaria and other parasites known as apicomplexans.
Results of the study will appear in the March 15 issue of Proceedings of the National Academy of Sciences. Scientists identified the new protein in the protozoan parasite Toxoplasma, which epidemiologists estimate infects 25 percent of all humans. The parasite rarely causes symptoms in most people, but can become a potentially life-threatening infection when the immune system is weakened by illnesses such as HIV or is suppressed to facilitate an organ transplant.
"Toxoplasma is like a time bomb that can go off and cause serious trouble when the immune system wanes or is compromised," explains L. David Sibley, Ph.D., professor of molecular microbiology at Washington University School of Medicine. For the study, researchers in Sibley's lab collaborated with Sinisa Urban, Ph.D., assistant professor of microbiology at Harvard University.
Sibley's lab studies Toxoplasma both because of the threat it poses to patients and for the potential insights it can offer into other apicomplexan parasites, which include malaria and Cryptosporidium. Malaria, which is spread by mosquito bites, kills at least 1 million people per year through damage to red blood cells and clogging of the capillaries that feed the brain and other organs. Cryptosporidium, which causes diarrhea, vomiting and other symptoms, is one of the most common causes of water-borne disease in the world.
"These other parasites are more devastating in terms of the number of people they affect, but they're somewhat harder to work on," Sibley explains. "Because it's so much easier to study, we use Toxoplasma as a way to ask about very basic things that occur in all of these parasites."
One area of enduring interest for Sibley and his colleagues has been the question of how Toxoplasma and other apicomplexan parasites move themselves around, given their lack of appendages or hairlike structures known as cilia or flagella. Researchers had determined that Toxoplasma has a rotating protein-based conveyor belt on its underside that enables it to move.
To push forward, any rotating object—be it a belt or a wheel—needs traction, or the ability to grip onto something and push against that grip. Car wheels, for example, are sometimes given added traction in wintertime via the addition of chains.
The parasite supplies its moving belt with traction by putting spots of an adhesive protein on it. The adhesive protein allows the host to attach to the exterior of the host cell. The resulting contact points give the belt something to push against as it moves backward, in turn pushing the parasite forward.
But these adhesive spots have a significant disadvantage in comparison to a chain: if the belt is to continue rotating, the glue spots have to be cut off at the back or they will jam the belt, greatly reducing the parasite's ability to move and thereby infect a host.
A postdoctoral researcher in Sibley's lab, Fabien Brossier, Ph.D., decided to try to use the recently completed Toxoplasma genome to search for the protein that lets the parasite detach glue spots at the back of the belt. Scientists led by the Institute for Genomic Research in Rockville, Md., posted the completed Toxoplasma genome online last year. Geneticists at Washington University's Genome Sequencing Center contributed to the effort.
Brossier knew the protein that did the work had to be a protease, a protein that cuts or degrades other proteins. He also knew it was likely to belong to a special subcategory known as rhomboid proteases, which are able to clip off sections of proteins near the surface of the cell membrane.
Using other rhomboid proteases as guides, Brossier identified genes for five rhomboid proteases in Toxoplasma. Scientists showed that the parasite used at least four of those genes to make active proteases. When they looked at where and when Toxoplasma made the proteases, they found that only one, TgROM5, consistently showed up at the back end of the parasite, where it would be ideally positioned to snip off the adhesive protein spots.
Cell tests in Urban's lab showed that only TgROM5 could cleave the proteins that make up the adhesive spots.
"There's a great deal of interest in this because TgROM5 might someday be targeted with inhibitors," Sibley says. "And by comparison to TgROM5 we can now point to highly similar proteins in malaria and other related protozoa that are also potentially promising targets for inhibitors."
Brossier F, Jewett TJ, Sibley LD, Urban S. A spatially localized rhomboid protease cleaves cell surface adhesins essential for invasion by Toxoplasma. Proceedings of the National Academy of Sciences, Vol. 102 (11), 4146-4151.
Funding from the International Human Frontier Science Program, the National Institutes of Health and the Burroughs Wellcome Fund supported this research.
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