Know more

From a tomato mutant producing fruits enriched in ascorbate (vitamin C), we identified a protein related to phototropins using mapping by sequencing. We then showed that this protein, named PAS/LOV and whose function remained unknown, is an inhibitor of an enzyme of the ascorbate synthesis pathway whose action is counteracted by blue light. This mechanism was demonstrated in vivo by cultivating plants under different wavelengths, then in vitro after purifying PAS/LOV and its target enzyme with heterologous expression systems. This discovery, which makes it possible to understand how blue light stimulates the synthesis of ascorbate in plants, opens the way to obtaining plants that produce more ascorbate, beneficial for human diet but also potentially more resistant to environmental constraints.

Context and challenges

Ascorbate, whose synthesis pathway in plants has only been known for 25 years (Wheeler et al. 1998), plays a major role as an antioxidant by participating in the elimination of reactive oxygen species usually produced in response to biotic and abiotic stresses but also under optimal conditions, in particular by photosynthesis (Decros et al., 2019). Due to its high antioxidant potential, ascorbate or vitamin C is one of the most important traits for the nutritional quality of fruits and vegetables. Indeed, evolution in humans and some animal species has led to the loss of the L-gulono-γ-lactone oxidase activity which catalyzes the last stages of its biosynthesis (Burns, 1957). Paradoxically, the domestication of various fruit species has led to a decrease in ascorbate content (Gest et al., 2013), suggesting the existence of a trade-off between yield and fruit quality. Thus, understanding ascorbate metabolism is a major challenge for plant breeding, especially for fleshy fruit species such as tomato, our main sources of vitamin C.

Results

A tomato mutant producing fruits and leaves enriched in ascorbate was detected and then studied (Bournonville et al; 2023). The causal mutation, identified by a mapping by sequencing strategy, affects PAS/LOV, a protein belonging to a class of photoreceptors. Its action as an inhibitor of ascorbate biosynthesis was demonstrated by gene editing with CRISPR/Cas9 that resulted in plants enriched in ascorbate. Functional characterization revealed that PAS/LOV interacts in the cytoplasm and in the nucleus with both isoforms of GDP-L-galactose phosphorylase (GGP), known as the most controlling step of the ascorbate synthesis pathway. An optogenetic approach then showed that blue light modulates this interaction. In vitro tests with heterologously expressed GGP and PAS/LOV finally showed that PAS/LOV is a non-competitive inhibitor of GGP and is inactivated by blue light.

Outlook

On a fundamental level, this discovery allows us to better understand how and why plants adjust the production of their main antioxidant to light intensity. On the applied level, PAS/LOV now represents a target for the selection of ascorbate-enriched plants, while blue light treatments could be tested to stimulate ascorbate synthesis. Increasing ascorbate production in a relevant way could ultimately stimulate plant defense and improve fruit preservation while increasing their nutritional quality.

By Yves Gibon & Pierre Baldet - Team Metabolism

References

• Bournonville C, Mori K, Deslous P, Decros G, Blomeier T, Mauxion J-P, Jorly J, Gadin S, Cassan C, Maucourt M, Just D, Brès C, Rothan C, Ferrand C, Fernandez-Lochu L, Bataille L, Miura K, Beven L, Zurbriggen MD, Pétriacq P, Gibon Y, Baldet P (2023) Blue light promotes ascorbate synthesis by deactivating the PAS/LOV photoreceptor that inhibits GDP-L-galactose phosphorylase ; The Plant Cell, in press, DOI : 10.1093/plcell/koad108
• Burns JJ (1957) Missing Step in Man, Monkey and Guinea Pig required for the Biosynthesis of L-Ascorbic Acid. Nature 180: 553-553. DOI:  10.1038/180553a0
• Decros, G., Baldet, P., Beauvoit, B., Stevens, R., Flandin, A., Colombie, S., Gibon, Y., Petriacq, P. (2019). Get the Balance Right: ROS Homeostasis and Redox Signalling in Fruit. Frontiers in Plant Science, 10, 1-16. DOI : 10.3389/fpls.2019.01091
• Gest N, Gautier H, Stevens R (2013) Ascorbate as seen through plant evolution: the rise of a successful molecule? Journal of Experimental Botany 64: 33–53. DOI : 10.1093/jxb/ers297
• Wheeler GL, Jones M, Smirnoff N (1998) Vitamin C in higher plants. Nature 393: 365-369. DOI:  10.1038/30728