For decades, the field of plant engineering has relied on Agrobacterium infection to transfer genes to plants. A widely used model organism, Nicotiana benthamiana, has become a workhorse in the plant biology field because of its unique ability to rapidly express genes delivered by Agrobacterium within just 2 to 5 days, sidestepping the lengthy process of waiting months to engineer a whole plant. This system allows researchers to quickly test and study the genetic transformations occurring in the plant.
This method has led to significant advances in basic plant biology research, and has also opened up the possibility of molecular farming, an industry focused on producing molecules on-demand within the leaves of a plant, allowing the plant to essentially act as a small factory.
“However, even though researchers have been using this system for decades, we have a very poor understanding of what’s happening under the hood,” said Patrick Shih, Director of Plant Biosystems Design at the Joint BioEnergy Institute (JBEI). “If we can learn more about what’s happening at a molecular level, we can use that knowledge to improve plant engineering.”
In a new study published in Nature Plants, JBEI researchers developed a modeling framework of the Agrobacterium system that provided insights into its molecular mechanisms. Using this framework, they were able to better understand the bottlenecks in the system and create solutions that ended up boosting the production of molecules in plants.
“For any sort of engineering application, you need to have a quantitative model of the system you’re working with. If you were trying to build a bridge or an airplane, you’d need to have a quantitative blueprint to guide you,” said Simon Alamos, a postdoctoral researcher at JBEI who led the study. “In plant engineering, Agrobacterium is like our workhorse, but we haven’t had a quantitative framework to model this system until now.”
Because the Agrobacterium system has worked well for plant engineering needs so far, it hasn’t been more thoroughly studied. But the field is rapidly becoming more technically complex.
“Twenty years ago, if you wanted to make a small molecule in plants, the metabolic pathways to produce those molecules were maybe three to five steps, and the Agrobacterium system worked quite well for this,” Shih said. “Today, these pathways are more complex — some are over 30 steps long. All of a sudden you start to realize there are technical bottlenecks in this system, and that if we understood this system better, we could more efficiently produce these molecules.”
The researchers found that Agrobacterium engages in biological interactions inside of the leaf that dictate how well genes transfer.
“We discovered that the bacteria can help each other, or they can compete with each other, which negatively affects their capacity to transform plant cells,” Alamos said. “They engage in these two kinds of interactions depending on the population density.”
Higher amounts of Agrobacterium in the leaf led to less efficient transformation of plant cells. The researchers sought to increase the number of genes that Agrobacterium can deliver without having to increase the population density.
To do that, they created a modification to the Agrobacterium system that allows the bacteria to carry two genes at a time, as opposed to one in the traditional system. Using this modification, called “BiBi,” the researchers demonstrated they could significantly improve the production of their target molecule. The researchers have filed a patent on this technology that they say could be transformative for the field of plant engineering.
This method could also aid in making biofuel production more viable. One of JBEI’s strategies centers around the idea that plants can be engineered to have a dual use — its plant biomass can be used for making biofuels, while the plant itself can produce valuable compounds within its leaves.
“That way, you get more bang for your buck from the plant that you’re growing,” Shih said. “Engineering plants so that we can generate commodity chemicals within them is going to help subsidize the cost of making a biofuel from that same plant’s biomass. This will help make the whole process more economically feasible.”
The Joint BioEnergy Institute is a DOE Bioenergy Research Center.