ANN ARBOR, Mich. -- Protected by a tough outer coat that is impervious to cold, heat, drought and harsh chemicals, anthrax spores can remain dormant in the soil for decades. Once inside a living host, however, they can germinate and begin infecting cells in as little as ten minutes.
Scientists know very little about what triggers an anthrax spore to break dormancy. Identifying the biochemical signals that start the process is an important first step to preventing anthrax infection.
A new study by University of Michigan scientists John A.W. Ireland, Ph.D., and Philip C. Hanna, Ph.D., shows that germination requires the coordinated activity of several genes, receptor proteins and amino acids in at least two simultaneous signaling pathways. The U-M study, published in the March 2002 issue of the Journal of Bacteriology, is the first to match anthrax genes with specific amino acids and signaling pathways that trigger germination.
"Anthrax doesn't rely on a single signal," says Hanna, an assistant professor of microbiology and immunology in the U-M Medical School. "Endospores have a redundant germination mechanism. It's the bug's way of ensuring that it won't lose its protective armor until conditions are right for germination."
Hanna and Ireland discovered that amino acids, the fundamental building blocks of all proteins in the body, in combination with purine ribonucleosides, the building blocks of DNA and RNA, are triggers for anthrax spore germination. The process appears to begin when receptor proteins on the spore's membrane bind to ring-shaped or aromatic structures on certain amino acids and ribonucleosides.
"The receptor protein is the lock and ring structures are the keys," says Ireland.
"The only place we know where all the required elements for germination are present is inside our cells, especially our phagocytes -- the scavenger cells of the immune system," Ireland explains. "But even in the macrophage where conditions are optimum for germination, the spore remains intact until at least two separate signaling pathways are activated."
Because it can be handled safely outside a high-level bio-containment laboratory, Hanna and Ireland used Sterne-based strains of anthrax in their research. The Sterne strain has been altered to remove dangerous infectious segments of the anthrax molecule called plasmids.
U-M scientists checked the genetic sequence of Sterne genes used in their study against the same genes in Bacillus anthracis Ames, the deadliest strain of anthrax, and found them to be a close match. B.anthracis gene sequences were provided by Tim Read, senior scientist at the Institute for Genomic Research (TIGR) in Rockville, Maryland.
Major findings of the study include:
Host signals from alanine and inosine were strong spore cogerminants when combined with other amino acids.
- Alanine was capable of triggering a strong, rapid germination response independently, but only when alanine concentrations were many times above normal cellular levels.
- Germinants vary depending on the anthrax bacterial species. Inosine is a strong independent germinant for B. cereus, but not for B. anthracis.
- A gene called gerS, present in both the Sterne strain and Ames strain of B.anthracis, appears to trigger two major signaling pathways. Similar genes have been found in other species of anthrax and may control germination in those species.
- Receptor proteins produced when the gerS gene is active respond specifically to the aromatic rings on certain types of amino acids and ribonucleosides.
In future research, Hanna will test these amino acids and ribonucleosides to see if they trigger anthrax spore germination in tissue cultures. Eventually, he hopes to expand the study to animal models. His research is supported by the National Institutes of Health and the Office of Naval Research.
Journal of Bacteriology, March 2002, p. 1296-1303, Vol. 184, No. 5
Editors: Full text of the published article is available on the ASM web site at: http://jb.asm.org/cgi/content/abstract/184/5/1296
Journal
Journal of Bacteriology