Vol 5. Issue 31 / October 17, 2005
The Face of Tumefaciens
By Jason Socrates Bardi
Agriculture has a long relationship with the bacterium Agrobacterium tumefaciens. Plants plagued with a pathogenic strain of this bacterium suffer a debilitating condition called crown gall disease, which is characterized by the growth of large "gall" tumors around the crowns of the roots that cannot be cured. Because some 600 different plant species are susceptible to crown gall disease, the bacterium is no small enemy to agriculture.
"It infects olives, cherries, apples, and a whole range of different plants," says John Reader, a former research associate at The Scripps Research Institute who is now an assistant professor at the University of North Carolina. At Scripps Research, Reader worked in the laboratory of Professor Paul Schimmel, where they both turned their attention to crown gall disease.
The thing that makes crown gall disease incurable is exactly what makes it of huge interest to agricultural science. The disease is incurable because of an unusual property of Agrobacterium tumefaciens: the bacterium has the ability to inject a fragment of its DNA into the nucleus of the plant cells it is infecting. Once inside, the bacterial DNA is integrated into the plant cell's genome and essentially becomes a permanent part of the cell.
This fact makes Agrobacterium tumefaciensis an agricultural pathogen of the first degree—one that defines the very word "pathos," the Greek root that means suffering. Though the foreign DNA is quite small, it will hijack the cell and take over the cell's growth, leading to unsightly tumors. These grow large and sap the energy of the plant. And as one tree suffers, so potentially do others. The spread of crown gall disease in an orchard of fruit-bearing trees can be devastating to crops, dramatically reducing the trees' yield.
Unless, that is, the trees are treated as seedlings with a bacterial biocontrol agent that is known to prevent crown gall disease through a mechanism that nobody has sufficiently explained until now.
In an article in a recent issue of the journal Science, Reader, Schimmel, and their colleagues have figured out how this bacterial biocontrol agent prevents Agrobacterium tumefaciens from growing. They collaborated in their chemical forensics with Stephen Farrand at the University of Illinois at Urbana–Champaign, an investigator who has had a long program of research studying Agrobacterium tumefaciens and the bacterial biological control agent.
Mistaking Poison for Food
A crown gall tumor is quite a remarkable thing. The plant cells in the tumor are not simply growing abnormally and contributing to the formation of the crown galls—they also become little factories that produce the chemicals the pathogenic Agrobacterium tumefaciens needs to feed on. Like an ecosystem, the infected plant will feed the very bacterium that infects it, and with all the live bacteria it supports, a single tree suffering from crown gall in an orchard can then spread the disease to other trees.
For years, orchard managers have been successfully controlling crown gall disease by treating seedlings with a closely related but non-pathogenic bacterial strain known as Agrobacteriumradiobacter strain K84. Seedlings placed in a compost containing this agent are resistant to crown gall disease.
This protection occurs because of a toxic trick that the K84 strain plays on the Agrobacterium tumefaciens. K84 produces a "Trojan horse" chemical toxin called agrocin 84 that looks almost identical one of the unique chemical substrates that an infected tree produces to feed Agrobacterium tumefaciens—called agrocinopine A.
Because of this molecular mimicry, the bacteria readily gobble down the chemicals, and once inside the bacteria, the agrocin 84 molecules are transformed into toxins. Thus the Agrobacterium tumefaciens eat the agrocin 84 chemicals and die—sort of like the Roman Emperor Claudius quaffing the poisoned mushrooms, which he loved to eat.
Why does the one bacterium pull an "I, Claudius" on the other? They are competing species. The Agrobacterium tumefaciens has a major advantage in that it can become entrenched within the plant itself, but the K84 bug is able to outcompete Agrobacterium tumefaciens by tricking it.
What Reader, Schimmel, and their colleagues figured out is how exactly the bacterial biological control agent works, which is something that had only been guessed at before. Because agrocin 84 has a core that looks like an adenine nucleoside, many scientists assumed that it would follow a mechanism somewhat like the AIDS drug AZT, a nucleoside analogue that is incorporated into a growing chain of DNA and halts the process known as transcription.
But, surprisingly, the scientists found that instead of blocking transcription, the toxin targets one of the critical enzyme-driven steps of protein translation—the charging of a tRNA with an amino acid by a tRNA "synthetase" enzyme.
Synthetase molecules are ubiquitous in nature. As we know them today, they are involved in one of the most fundamental processes in life—translation, which is the culminating step in the expression of a gene whereby nucleotide bases are translated into amino acid proteins. Synthetase molecules play a crucial role in this because they charge individual tRNAs with the correct amino acids. "Life cannot be sustained without them," says Schimmel.
In the process of attachment of amino acids to tRNA molecules, one of 20 different synthetases binds to its corresponding amino acid, a molecule of ATP, and one of 20 different tRNA molecules. The synthetase then hydrolyzes the ATP, attaches the amino acid to the end of the tRNA, and lets go.
By looking at the chemical structure of the toxin agrocin 84, Reader, Schimmel, and their colleagues realized that it looks similar to a catalytic intermediate in the charging of leucyl tRNA with a leucine amino acid by a leucyl-aminoacyl tRNA synthetase.
"It was striking to us that it looked so similar," says Schimmel.
Agrocin 84 is effectively an inhibitor of the synthetase, and it prevents the formation of aminoacylated tRNA. This gums up protein synthesis in the bacterium, and leads to the death of Agrobacterium tumefaciens.
Interestingly, the K84 bug carries a second leucyl-tRNA synthetase that is resistant to the toxin. This self-protective enzyme allows it to survive its own toxic onslaught.
The article, "Major Biocontrol of Plant Tumors Targets tRNA Synthetase" by John S. Reader, Phillip T. Ordoukhanian, Jung-Gun Kim, Valerie de Crécy-Lagard, Ingyu Hwang, Stephen Farrand, and Paul Schimmel appears in the September 2, 2005 issue of the journal Science. See: http://dx.doi.org/10.1126/science.1116841
This work was supported by the National Institutes of Health, a National Foundation for Cancer Research fellowship, a grant from the Crop Functional Genomics Center, and support from the Skaggs Institute for Chemical Biology at The Scripps Research Institute.
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