Scientists at the Earlham Institute in Norwich, U.K., say they have been able to turn tobacco plants into solar-powered factories for moth sex pheromones. In addition, they’ve shown how the production of these molecules can be efficiently managed so as not to hamper normal plant growth.

Farmers can hang pheromone dispersers among their crops to mimic the signals of female insects, trapping or distracting males from finding a mate. Some of these molecules can be produced by chemical processes but chemical synthesis is often expensive and creates toxic byproducts.

Nicola Patron, PhD, who led this new research and heads the synthetic biology group at Earlham, believes the team’s work will pave the way to routinely using plants to produce a wide range of valuable natural products.

Scientists engineered a species of tobacco, Nicotiana benthamiana, to produce moth sex pheromones. [Martin Barraud/Getty Images]
“Synthetic biology can allow us to engineer plants to make a lot more of something they already produced, or we can provide the genetic instructions that allow them to build new biological molecules, such as medicines or these pheromones,” said Patron.

In this latest study “Tunable control of insect pheromone biosynthesis in Nicotiana benthamiana”, which appears in Plant Biotechnology, the researchers worked with scientists at the Plant Molecular and Cell Biology Institute in Valencia to engineer a species of tobacco, Nicotiana benthamiana, to produce moth sex pheromones. The same plant has previously been engineered to produce Ebola antibodies and even coronavirus-like particles for use in COVID vaccines.

New DNA sequences

The group built new sequences of DNA in the lab to mimic the moth genes and introduced a few molecular switches to precisely regulate their expression, which effectively turns the manufacturing process on and off.

“Previous work has demonstrated that plants can be used as production platforms for molecules used in health, medicine, and agriculture. Production has been exemplified in both stable transgenic plants and using transient expression strategies. In particular, species of Nicotiana have been engineered to produce a range of useful molecules, including insect sex pheromones, which are valued for species-specific control of agricultural pests,” write the investigators.

“To date, most studies have relied on strong constitutive expression of all pathway genes. However, work in microbes has demonstrated that yields can be improved by controlling and balancing gene expression. Synthetic regulatory elements that provide control over the timing and levels of gene expression are therefore useful for maximizing yields from heterologous biosynthetic pathways.

“In this study, we demonstrate the use of pathway engineering and synthetic genetic elements for controlling the timing and levels of production of Lepidopteran sex pheromones in Nicotiana benthamiana. We demonstrate that copper can be used as a low-cost molecule for tightly regulated inducible expression. Further, we show how construct architecture influences relative gene expression and, consequently, product yields in multi-gene constructs.

“We compare a number of synthetic orthogonal regulatory elements and demonstrate maximal yields from constructs in which expression is mediated by dCas9-based synthetic transcriptional activators. The approaches demonstrated here provide new insights into the heterologous reconstruction of metabolic pathways in plants.”

Fine tuning pheromone production

An important component of the new research was the ability to fine tune the production of the pheromones, as coercing plants to continuously build these molecules has its drawbacks.

“As we increase efficiency, too much energy is diverted away from normal growth and development,” explained Patron. The plants are producing a lot of pheromone but they’re not able to grow large, which essentially reduces the capacity of our production line. Our new research provides a way to regulate gene expression with much more subtlety.

“We’ve shown we can control the levels of expression of each gene relative to the others. This allows us to control the ratio of products that are made.

“Getting that recipe right is particularly important for moth pheromones as they’re often a blend of two or three molecules in specific ratios. Our collaborators in Spain are now extracting the plant-made pheromones and testing them in dispensers to see how well they compare to female moths.”

A major advantage of using plants is that it can be far more expensive to build complex molecules using chemical processes, noted Patron. Plants produce an array of useful molecules already so novel techniques to adapt and refine the existing machinery.

“In the future, we may see greenhouses full of plant factories providing a greener, cheaper and more sustainable way to manufacture complex molecules,” she pointed out.

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