"This work is important because pathogenic bacteria such as Pseudomonas aeruginosa use protein secretion systems to cause disease in their hosts," said Joseph Mougous, lead author of the study published in the June 9 issue of Science. "In the case of Pseudomonas aeruginosa, the host may be a cancer patient with a weakened immune system, a burn patient, or a person with cystic fibrosis (CF)."
Pseudomonas aeruginosa (PA), a pathogen that infects more than 80 percent of cystic fibrosis patients, is a leading cause of these patients' death. PA is difficult to treat because it is resistant to many drugs.
"Since we know so little about what this bacterium is up to while it's engaged in these chronic infections, the discovery of this protein secretion system might lead to finding a new target for treatments," said Mougous, a research fellow in the Harvard Medical School (HMS) Department of Microbiology and Molecular Genetics.
Researchers at Argonne National Laboratory (ANL) provided one of the clues that contributed to the HMS discovery. Working through a number of pathogenic proteins, ANL protein crystallographer Marianne Cuff saw a bagel-shaped pore that might be involved in transferring toxins into cells. She deposited the structure of the protein, called Hcp1, into the Protein Data Bank, a resource used by biologists worldwide to find information about the proteins they are studying.
While exploring the Protein Data Bank, Mougous, who was studying PA in the laboratory of department chair John Mekalanos, recognized that the amino acid sequence of Hcp1 in PA closely resembled that of Hcp1 in Vibrio cholerae. The Mekalanos lab had previously discovered that the Hcp1 protein of V. cholerae is released from the bacterium via a novel secretion pathway. Because Hcp1 proteins from both pathogens belong to the same protein family, Mougous wondered whether the Pseudomonas Hcp1 might also be secreted via this pathway.
The Harvard and Argonne researchers quickly formed a collaboration and confirmed the hypothesis. They then turned their attention to Hcp1 in cystic fibrosis patients to gain more insight in the role of Hcp1 during infection.
Working with cystic fibrosis patients at Children's Hospital Boston, the HMS researchers sought and found Hcp1 in the sputum of patients with PA. They also found Hcp1 antibodies in the patients' blood - further evidence that Hcp1 plays a critical role in the infection. The human immune system creates antibodies to pathogens it is exposed to.
"CF patients are particularly susceptible to PA," Mougous said. "The bacterium thrives in the excess mucus that accumulates in their lungs. Once a PA infection in a CF patient's lung has been established, these hardy bacteria are difficult or impossible to clear, which over many years eventually results in the death of the patient. Our paper provides evidence that the protein secretion system we discovered represents at least one way this bacterium could be promoting its own survival in the lungs of CF patients."
"This finding provides a possible drug target to fight the infection in cystic fibrosis patients," added Andzrej Joachimiak, director of Argonne's Structural Biology Center and of the Midwest Center for Structural Genomics based at Argonne.
The research was supported in part by grants from the National Institutes of Health, the Damon Runyon Cancer Research Foundation, and the U.S. Department of Energy, Office of Biological and Environmental Research.
Harvard Medical School