Demonstrating that nature is no redder in tooth and claw than is industry, scientists have imposed a ruthless scheme of negative selection upon genetically engineered bacteria. The scheme effectively culls laggards from the microbial herd, leaving it, in a sense, fitter—so much fitter, in fact, that surviving bacteria show up to 30-fold increases in chemical output.
Enriching populations of genetically engineered bacteria for rare “high producers” can be a tedious business, requiring the attention of bioengineers, who often find it necessary to construct and evaluate many alternative designs. To avoid all this trouble, researchers at the Wyss Institute led by geneticist George Church, Ph.D., decided to let bacteria compete with each other, and thereby allow the high producers to emerge without direct human oversight. With bacterial self-monitoring, production can be speeded nearly 1,000-fold.
The trick, however, was to control for unproductive cheaters, which tend to proliferate in schemes of artificial selection. To minimize the carryover of cheaters while preserving bacterial diversity, the Wyss researchers used sensory proteins responsive to a number of target chemicals to couple the concentration of the target chemical in each cell to individual cell fitness.
The details of this approach appeared December 16 in the Proceedings of the National Academy of Sciences, in an article entitled, “Evolution-guided optimization of biosynthetic pathways.”
“We report a general strategy that combines targeted genome-wide mutagenesis to generate pathway variants with evolution to enrich for rare high producers,” wrote the authors. “We convert the intracellular presence of the target chemical into a fitness advantage for the cell by using a sensor domain responsive to the chemical to control a reporter gene necessary for survival under selective conditions.”
“We make the bacteria addicted to the chemicals we want them to produce,” said Jameson Rogers, a lead co-author of the study, Ph.D. candidate at Harvard School of Engineering and Applied Science and Wyss Institute graduate researcher. “Then, we treat them with an antibiotic that only allows the most productive cells to survive and make it on to the next round of evolution.”
The technique makes a desired chemical product essential to the bacteria's survival by modifying their DNA so that antibiotic–resistant genes are activated, but only in the presence of a certain chemical, such as the one that is desired for production. At the same time, the genetic modification makes the low-output chemical producers highly susceptible to being killed off by antibiotics. Only the most productive cells generate enough of the desired chemical to be completely resistant to the antibiotic and survive to go onto the next round of evolution. As each evolution cycle progresses, the bacteria become more and more effective at producing the desired chemical as they use the “survival of the fittest” principle to stamp out the weakest producer cells.
“We're using evolution to select for the cells that only serve our purpose best, making human monitoring less important to that feedback loop and instead relying on the bacteria to self-monitor their production performance,” said Dr. Church, the study's senior author, who is a Wyss Institute Core Faculty member, Professor of Genetics at Harvard Medical School and Professor of Health Sciences and Technology at Harvard and MIT. “This is a major direction of growth in synthetic biology, where the focus has mostly been on one-off experiments until this point.”