Comprehensive analysis of single plant cells provides new insights into natural product biosynthesis, a University of Georgia study shows

Single-cell multi-omics reveals that cell types are differentially involved in the production and accumulation of medically relevant plant compounds.

Plants are impressive in their diversity, but especially in the variety of metabolites they produce.

Many plant-natural products are highly complex molecules, such as the alkaloids vincristine and vinblastine, which are produced by the Madagascar periwinkle Catharanthus roseus. These two substances are already indispensable in cancer therapy.

Researchers are very interested in finding out which individual biosynthetic steps are required to form complex molecules.

First author Chenxin Li, of the University of Georgia’s Center for Applied Genetic Technologies, said: “Currently, these compounds are still obtained in very small quantities from the plant’s leaf extract.

“We can learn from the plant how this compound is produced and use this knowledge to develop production systems that are more cost-effective, scalable and sustainable.”

Scientists know that gene activity is not the same in all cells of a plant and that the chemistry can differ drastically from cell to cell.

Therefore, the goal of the current study was to use a new set of methods collectively termed ‘single-cell omics’ to investigate specialised and rare cell types that play a central role in the biosynthesis of plant natural products, and whose signals are often obscured by more abundant cell types in plant organs.

One of the lead authors, Lorenzo Caputi, head of the Alkaloid Biosynthesis Project Group in the Department of Natural Product Biosynthesis in Jena, said: “With single-cell omics, we have a method that allows researchers to assign genetic and metabolic information to individual cells.

“The term ‘omics’ refers to the fact that an entire collection of genes or metabolites is quantified and analysed.”



Biosynthetic pathway of vinblastine


As the analyses showed, the entire biosynthetic pathway for the alkaloid vinblastine is organised in three stages and three discrete cell types.

Li said: “The first stage is expressed exclusively in specialised cells associated with vascular bundles in the leaf, called IPAP.

“The second stage of the biosynthetic pathway is expressed only in cells of the epidermis, the layer of cells that cover the leaves.

“The last known steps of the biosynthetic pathway are expressed exclusively in idioblasts, a rare cell type of the leaf.”

The researchers measured the concentrations of several intermediates in the metabolic pathway for vinblastine in single cells and were surprised.

Sarah O’Connor, head of the Department of Natural Product Biosynthesis at Jena, said: “Two important precursors of vinblastine, catharanthine and vindoline, occur in the idioblast cells at millimolar concentrations, about three orders of magnitude higher than vinblastine itself.

“The concentration of the two precursors in these cells was much higher than we expected and even exceeded their concentrations in whole organ extracts.

“However, this observation makes sense in that catharanthine and vindoline were found only in the rare idioblast cells.

“The abundant other cells in the leaf dilute the high concentration when whole leaves are crushed.”


Natural products


The research team is confident that the organisation of biosynthetic pathways for medicinally relevant alkaloids in Catharanthus roseus is not an isolated phenomenon.

Robin Buell, Professor at Georgia University, said: “We are just beginning to understand how and why such a cell type-specific organisation exists.

“In addition, analysis of genes expressed simultaneously in a particular cell type has helped us identify new players in this metabolic pathway. The same technique can be used to study the biosynthesis of many other natural products.

“Finally, the exact sites of accumulation of plant compounds, such as the epidermis, the vascular system, or latex duct, can help us hypothesise the ecological roles of natural products.

“For example, depending on the pattern of accumulation, the compounds may be more effective against biting insects than they are against sap-sucking insects.”

A better understanding of the biosynthetic pathways of the anti-cancer drugs vincristine and vinblastine may also help to produce or harvest these compounds more effectively in the long term.

The use of the methods described is also promising for the study of many other interesting and medically important natural products from the plant kingdom.

The approach described here will help to narrow down these rare and specialised cells and uncover the gene activities and chemistry that are exclusive to them.

Image: The Madagascar periwinkle (Catharanthus roseus) of the dogbane family produces a number of alkaloids of medicinal interest. Analyses at the level of different cell types enabled the discovery of still missing genes for the biosynthesis of the two most important natural products from the plant, vincristine and vinblastine (skeletal formula), which are used as chemotherapeutic agents in the treatment against cancer. Credit: Angela Overmeyer, Max Planck Institute for Chemical Ecology (CC BY-SA).

Research Aether / Earth Uncovered