Anthrax breakthrough reported
Researchers at the Medical School McArdle Laboratory for Cancer Research and their collaborators at Harvard Medical School have found the receptor — a docking structure — that anthrax toxin binds to in order to enter cells.
The finding, to appear in the Thursday, Nov. 8, Nature, represents a major advance in understanding exactly how the toxin kills host cells, leading quickly to death. The structure consists of a single protein the researchers call anthrax toxin receptor (ATR).
By genetically engineering a portion of ATR, the scientists have also produced a form of it that can block the toxin from entering cells, a feat that may have crucial implications for future approaches aimed at treating anthrax infection.
“Our short-term goals are to study the mechanism of toxin uptake through ATR and to make enough of the toxin-blocking form of the receptor so that it can be tested in animal systems,” says senior author John A. T. Young, the Howard M. Temin Professor of Cancer Research. “A more long-term application would be for pharmaceutical companies to use the receptor along with anthrax toxin to screen the millions of compounds they’ve already synthesized to identify toxin inhibitors.”
Once it enters its host, the anthrax bacterium secretes a toxin consisting of three components. Two of them — edema factor (EF) and lethal factor (LF) — wreak havoc inside cells. The third component, protective antigen (PA), binds to ATR and acts like a doorway for EF and LF to enter the cell.
In earlier studies, Kenneth Bradley, Young’s graduate student, mutated cells to find those that were resistant to infection caused by retroviruses, the family of viruses that includes HIV. His purpose was to identify unknown cellular proteins that aid in virus production.
Bradley found that the cells had become resistant because they no longer produced surface virus receptors, proteins that were already known. Realizing that this method could be a powerful approach for isolating other unknown cell-surface receptors, Bradley then applied a similar process to identifying the anthrax toxin receptor by first looking for cells that were resistant to toxin action.
“Initially we didn’t know the resistant cells had lost receptors. All we knew was that they could not be killed by components of the anthrax toxin,” says Young, noting that whole anthrax bacteria were not used in the studies.
In collaboration with Jeremy Mogridge, a post-doctoral fellow in the laboratory of John Collier at Harvard Medical School, Bradley then showed that the cells had in fact lost the receptor for anthrax toxin. To find the receptor, Bradley introduced a collection of genes into the cells and found a single gene, ATR, that restored receptor production.
The researchers analyzed the gene in detail and compared it to existing genes. They found ATR’s make-up to be related to that of TEM8, a gene product produced at higher than normal levels in people with colorectal cancer.
ATR was also found to include one region that protrudes outside the cell membrane, away from the cell interior. The researchers showed that this region of ATR is the precise docking site to which PA adheres. Additional experiments performed by Mogridge and Michael Mourez of the Collier lab showed a direct and specific interaction between the special ATR region and the receptor-binding area of PA.
By extracting the special region from the rest of ATR and releasing it into the extracellular area, the scientists found they could entice PA away from the cell surface. This action prevented PA from entering the cell and from delivering a specifically engineered toxin.
Bradley says more studies examining the link between the colon cancer gene and ATR were likely. “We hope to be able to understand the normal function of ATR, which may shed new insights into cancer as well as anthrax toxin action.”
The Wisconsin Alumni Research Foundation and Harvard Medical School have filed a joint patent on the anthrax toxin receptor.