Understanding how protein toxins interact with cellular processes is crucial for developing more effective treatments for affected patients. Frederic Bard at the A*STAR Institute of Molecular and Cell Biology and co-workers have now used a genetic screening procedure to identify potential target molecules that could be used as antidotes for the problematic protein toxins ricin and Pseudomonas exotoxin.
Ricin is found in beans (pictured) of the plant Ricinus communis, which is used for castor oil production and relatively easy to obtain and prepare, as well as being stable when purified and dried. However, it is also potentially lethal when inhaled, even in tiny amounts. Pseudomonas exotoxin, on the other hand, is produced by the medically important bacterium Pseudomonas aeruginosa. The species is an opportunistic pathogen that often attacks burn wounds and weakened immune systems, and is a common lung infection in cystic fibrosis patients. Once an infection has started, the bacterium secretes the exotoxin, which damages cells and tissues.
Like many other toxins, ricin and Pseudomonas exotoxin disrupt protein synthesis. To achieve this disruption, however, they must first reach the inner core of cells by following the retrograde membrane traffic pathway—a complex route inside the cells. “We wish to understand how the toxins can hijack the retrograde pathway,” explains Bard. “We knew from previous studies that these proteins need components of this pathway, but the extent of this has been unclear.”
The researchers searched for host molecules required by both toxins that enable them to enter cells. Bard and his team aimed to block the expression of host genes individually and to see whether the toxins could still enter the cells in the absence of the proteins encoded by these genes. To achieve this on a large scale, they used RNA interference technology to screen the human genome efficiently for the necessary genes.
“We showed that each toxin requires several hundred host molecules, but that the actual amount of overlap between both sets of required genes is quite modest,” says Bard. Despite this, many of the genes shown by Bard’s team to be important gene-encoding proteins in membrane traffic are also found in similar cell compartments and structures.
“Our findings suggest that ricin and exotoxin exploit two intertwined retrograde pathways that converge and diverge at multiple points,” says Bard. The researchers hope that their work will contribute to the development of effective toxin antidotes and therapeutic treatments with limited side effects.
The A*STAR-affiliated researchers contributing to this research are from the Institute of Molecular and Cell Biology.