STANFORD, Calif. - The unusual ability of the organism Toxoplasma to infect and reproduce inside almost all warm-blooded animals has led scientists to wonder about the tricks it uses so successfully to subvert the behavior of cells. Now, a team of Stanford University School of Medicine researchers, led by John Boothroyd, PhD, has shown for the first time how Toxoplasma manages to be so effective: They documented how it injects a particular protein into the cell it infects and how that protein then travels to the cell's nucleus - where it blocks the cell's normal response to invasion.
Never before have researchers offered such insight into the way this type of parasite can hijack a host cell's genetic machinery for its own benefit. And the discovery has wide-ranging implications for a number of diseases caused by other parasites in this class, which reproduce only inside of cells, including the parasite that causes malaria.
The results will be published in the Dec. 20 edition of Nature. They come on the heels of another paper from Boothroyd's lab, published earlier in December in the journal Science, identifying two proteins that can determine how much damage the parasite Toxoplasma can inflict on an animal. Boothroyd is a professor of microbiology and immunology at the School of Medicine.
The latest findings reveal a new mechanism for how an intracellular pathogen can interact with its host, and they may help to explain important differences in how various Toxoplasma strains have evolved to exploit this interaction, said Susan Coller, PhD, one of the study's lead authors who was a postdoctoral scholar in Boothroyd's lab when the work was done.
What shocked the researchers was that a single protein was responsible for the dramatic differences between the strains; they had expected it to be much more complex.
"That it travels to the host cell nucleus is the cherry on the sundae," Coller said. "It's the heart of the cell, the ultimate prize. If you want to affect the cell in a dramatic way, go straight there."
The researchers found that Toxoplasma injects a protein called ROP16 into the host cell. ROP16 is a class of enzyme called a kinase, which is a mediator of cellular messages. Kinases are used by all cells to regulate a variety of key physiological processes, including responding to the presence of an invader. Injecting kinases is an extremely efficient way for a parasite to co-opt a host cell for its own purposes, Boothroyd said.
According to the study, different forms of the injected kinase have dramatically different effects on how a host cell responds to the invading parasite. Knowing what determines the extent of the immune response may allow for therapeutic manipulations, perhaps leading to physicians being able to tune down a response that's out of control in some cases of toxoplasmosis. Although Toxoplasma infections in humans are often asymptomatic, they can cause severe problems in isolated cases, particularly for individuals with compromised immune systems and for fetuses.
In North America and Europe, there are three main strains of Toxoplasma. Experiments have shown that the effects on mice infected with Toxoplasma are highly dependent on the type of strain. Recent results indicate that differences in infection might exist in humans too.
"When you look at the three different strains under the microscope, you can't distinguish them, yet they have such different properties," said the article's other lead author, Jeroen Saeij, PhD, a postdoctoral scholar in Boothroyd's laboratory. "Trying to find which parasite genes are responsible is like solving a puzzle."
The researchers sought to test the hypothesis that some of the strain-specific differences are a result of how the strains interact with the host cell. To do this, the researchers looked for large changes in the gene expression of the hosts - in this case, human cells - when they became infected.
The team used microarrays to examine the entire human genome's response to infection of cells with Toxoplasma. They pinpointed a number of genes involved in the immune response that were activated after being exposed to the parasite. Then, through a series of logical assumptions, they identified the Toxoplasma protein ROP16 as the culprit for causing the immunological changes in human cells. A key point is that it was responsible for the strain-specific differences in how the host cell responded to infection.
Each version of the ROP16 gene evolved to tweak the cells of a given host to varying degrees, Boothroyd said. "We hypothesize that, depending on which version of the ROP16 gene a given strain carries and which host is infected, it may carry out this task with greater or lesser efficiency," he said. "As a result, when a strain infects that host, the 'tweaking' is just right and the host is successfully infected with the minimum of damage."
Yet in a different host infected by that same strain, the activity of the ROP16 protein may be too strong, causing the parasite infection to rapidly overwhelm and kill the host. Alternatively, that version of ROP16 may not work in a given host at all, causing an excessive immune response in the host.
"Obviously the organism needs some powerful tools to manipulate the host's immune system to ensure its survival," said Saeij. "So it is very well possible that each time Toxo encountered new hosts, it expanded its arsenal of tools (duplicating or evolving existing kinases) to deal with the new challenges."
This work involved a collaboration with Montana State University and was supported by grants from the National Institutes of Health, the Ellison Medical Foundation and the University of California University-wide AIDS Research Program. Postdoctoral scholar Jon Boyle, PhD, is the other Stanford researcher included as an author of this publication.
Stanford University Medical Center integrates research, medical education and patient care at its three institutions - Stanford University School of Medicine, Stanford Hospital & Clinics and Lucile Packard Children's Hospital at Stanford. For more information, please visit the Web site of the medical center's Office of Communication & Public Affairs at http://mednews.stanford.edu.
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