Plants are known to produce valuable molecules, many of which are hard to chemically synthesise. Engineering biology approaches at the Earlham Institute are helping to decode these biosynthetic pathways.
Our relationship with the plant kingdom is an old one, providing us with food, shelter, and plenty of inspiration.
Ancient healers, witchdoctors, and herbalists all experimented with the flora growing around them. More nefarious actors took a keen interest in the toxins, but a great many natural medicines were discovered.
Today, we know the bioactive molecules responsible for these harming or healing effects. And we’ve even managed to biosynthesise a few of them in the lab.
During the 20th century, synthetic chemistry revolutionised drug discovery and production, giving us analgesics, arrhythmia treatments, and even anti-cancer drugs. But the manufacture of some bioactives can be complex, expensive, and often produces a range of waste products.
In contrast, the plants - some of which you’ll find in the average back garden - are happily making them with the modest inputs of sunlight and water.
At the Earlham Institute, we’re applying the latest engineering biology approaches to see if a return to plants might be the future of sustainable bioactive production.
Dr Melissa Salmon is a Postdoctoral Scientist in the Patron Group. Her research aims to map out each of the steps needed for plants to make medicinal bioactives, along with understanding the genes involved in regulating production.
Current wisdom suggests these bioactives are manufactured and stored in trichomes - tiny finger-like projections that can make the stem or leaf surface appear hairy.
Melissa’s recent work has focused on the UK native Asteraceae family of flowers, which includes daisies and marigolds.
“These plants have been used for centuries to make infusions,” explains Melissa. “We now know they contain metabolites that are part of a family of molecules called sesquiterpene lactones.
“We find these at quite small concentrations in the leaves and flowers of the plants, but they have strong anti-inflammatory properties.”
Inflammation is linked to a range of health conditions, which means anti-inflammatory compounds are in high demand.
But plants have a habit of producing a multitude of metabolites, some of which may be toxic to humans. This means the first challenge is to identify the specific bioactive molecule of interest, and then map out the genes responsible for synthesising it.
“Extracts from these plants have been shown to have strong anti-inflammatory activity, and this has been linked to sesquiterpene lactones,” explains Melissa. “We now want to link the bioactivity to a specific molecule and decode the genes involved in its biosynthesis.”
Microscopy of the underside of a leaf showing non-glandular trichome hairs highlighted in turquoise, and glandular trichomes highlighted in pink.
Daria Golubova, a PhD student in the Patron Group, has been carrying out cytotoxicity and anti-inflammatory bioassays from extracts and pure compounds to look for these properties in plant extracts.
These will help to confirm the sesquiterpene lactones as having the desired anti-inflammatory properties. Knowing the structure of the molecule from existing studies, they have turned their attention to the genes involved in manufacturing it.
To do this, Melissa has called upon the expertise of the Technical Genomics Group at the Institute, who have helped to generate the transcriptomes.
This has involved both short- and long-read sequencing - using the latest platforms at the Earlham Institute - and core bioinformatics support for the assembly and annotation.
The annotated transcriptomes help the group to identify the candidate genes involved in synthesising the sesquiterpene lactones.
“We know quite a lot about the enzyme families involved in making these molecules, which means we can break down the synthesis into different steps,” explains Melissa.
“Each synthesis step is catalysed by one of these enzymes, so we can use knowledge of the enzyme families coupled with phylogenetics to pick out candidate genes from the transcriptome.”
To confirm each of these steps, Melissa uses a plant-expression system, which uses Agrobacterium - a natural pathogen of plants - to introduce the candidate gene DNA into Nicotiana benthamiana through leaf infiltration.
Once the candidate gene is present in the cells of N. benthamiana, the plant can use its existing molecular machinery and resources to express the gene.
“We can then extract the products directly from the leaves to see whether the candidate gene is making the product we predicted,” says Melissa.
This process can then be repeated for each step of the pathway to identify all the genes involved in building the sesquiterpene lactones.
Another strand of the group’s research is the role of copy number on plant metabolite evolution.
“We think a higher copy number gives these plants an advantage when they need to adapt to change,” explains Melissa.
“A lot of plants have multiple copies of the genome in their cells. This allows them to diversify their metabolite profiles, or even make more of a particular metabolite - essentially by simultaneously expressing more of the genes involved in synthesis.”
The level of adaptation is fairly limited. These metabolites are of little use when coping with a changing climate, for example. But the ability to tweak the properties of some metabolites could improve their chances of fighting off pests or attracting pollinators.
This is an area the group is exploring with their Decoding Biodiversity programme partners at the Royal Botanic Gardens, Kew.
They’re particularly interested in yarrow, Achillea millefolium, which is part of the Asteraceae family and another plant that produces sesquiterpene lactones.
“We’re investigating whether different ploidy levels - the number of genomes within a cell - can influence the types and amounts of certain molecules in yarrow, to identify the optimum sources of bioactive molecules,” says Dr Melanie-Jayne Howes, Senior Research Leader in Biological Chemistry and Initiative Leader for Biointeractions and Bioactive Molecules at the Royal Botanic Gardens, Kew.
“By examining the evolutionary relationships between different Achillea species through sequencing their DNA, we hope to determine if more closely-related species produce similar types of molecules and explore whether predictive approaches can be applied to identify which other Achillea species could be potential sources of bioactive molecules.
“This research is greatly enhanced by the analysis of specimens in Kew’s extensive plant collections, including in the Herbarium and the Living Collection of plants.”
The joint approach should provide new insights into the origins of novel chemodiversity in these plants, as well as linking back to Earlham Institute’s interest in developing routes to access and manufacture bioactives.
Melissa’s recent work has focused on the UK native asteraceae family of flowers, which includes Calendula Officinalis, Eupatorium cannabinum, and Achillea millefolium.
By teasing apart the different steps, and all the molecular actors involved, Melissa is ultimately hoping to find ways to precisely enhance the bioactives production pipeline.
One of the routes to achieving this involves looking back in evolutionary time to see how these plants developed the ability to produce the target metabolites.
“If we know the genes involved, we can study how those genes have evolved,” she explains. “We then look at similar plants to compare the genomes, which helps us to track the evolution of any genetic diversity.”
The tools needed to compare the genomes will be another key output from the Decoding Biodiveristy research programme.
The naturally occurring diversity they find may reveal avenues for genetic fine-tuning that could be used to increase the amount of the metabolite produced, improve efficiency, or to remove any toxic by-products coexisting in the plant.
“We still have some way to go but it’s such an exciting programme to be part of because we can clearly see the enormous potential of this work,” says Dr Nicola Patron, group leader at the Earlham Institute and Associate Professor in the Department of Plant Science at the University of Cambridge.
“That ability to both protect and benefit from nature must be at the core of any solutions to the urgent global challenges we face.”
