Plants and fungi exchange 'molecular messages' to establish beneficial symbioses

In a recent study published in New Phytologist, researchers from CRAG, in collaboration with the University of Torino, reveal a novel mechanism of cross-kingdom communication, providing new insights into how plants and fungi interact and live together with reciprocal benefits. The work, co-led by Ignacio Rubio-Somoza, CSIC researcher at CRAG, and Professor Luisa Lanfranco from the University of Torino, presents the first experimental evidence that fungal small RNA molecules (sRNAs) can silence plant genes to facilitate the establishment of a symbiotic relationship.

Ancient symbiosis for old and modern challenges

Approximately 450 million years ago, as plants began colonizing land, they encountered a harsh environment. Symbiotic partnerships between plants and fungi, particularly mycorrhizal fungi, became crucial for survival. These partnerships, based on a trade of nutrients, allow plants to obtain mineral nutrients such as phosphorus, one of the main limiting factors for plants growth, while fungi depend on the carbon supplied by the host plant.

Nowadays, we know that mycorrhization directly boosts not only plant mineral nutrition but also the natural capacity of plants to cope with biotic and abiotic stresses, as well as the photosynthetic capability and therefore CO2 fixation. Most land plants, including many crops, engage in this symbiosis in both natural and agricultural systems.

Arbuscular mycorrhizal (AM) symbiosis, one of the most ancient and widespread forms of this relationship, involves nutrient exchange at specialized, tree-like fungal structures called arbuscules, which are formed inside root cortical cells of plants. The formation of AM symbiosis requires tightly regulated molecular communication, ensuring mutual recognition and cooperation between both organisms.

Gene silencing across species

A key step for granting establishment of AM symbiosis is that plants recognize beneficial fungi as friends, and they don’t activate defense mechanisms as they would be a threat. That recognition is based on the molecular dialogue between both organisms. Current research shows that sRNAs are essential elements in this process, by regulating gene expression through gene silencing. Cross-kingdom RNA interference (ckRNAi) had been previously described as a key mechanism in plant-pathogen interactions in works pioneered by the co-author of this publication, Hailing Jin, from UC Riverside.

This new study published in New Phytologist demonstrates, for the first time, that sRNAs from the AM fungus Rhizophagus irregularis play a critical role in establishing this symbiosis. Specifically, a fungal sRNA named Rir2216 was found to hijack the RNAi machinery of the Medicago truncatula to silence the plant transcription factor MtWRKY69, leading to a promotion of fungal colonization.

Using advanced techniques like laser microdissection, researchers specifically analyzed arbuscule-containing cells and found that MtWRKY69 expression was downregulated compared to non-mycorrhizal cells. Molecular biology experiments confirmed that Rir2216 reduces MtWRKY69 levels and functions as a key player in ckRNAi.

″These micro-messages could be the ‘Esperanto’ between different organisms, as different as belonging to different kingdoms″, Ignacio Rubio-Somoza points out.

In addition, researchers found that plants with increased levels of MtWRKY69 expression showed a reduced mycorrhization, with a lower percentage of frequency, intensity and arbuscule abundance. This indicates that regulation of MtWRKY69 expression levels plays a role in controlling the extent of fungal colonization in mycorrhizal roots, confirming the biological significance of the finding.

Next steps and applications in agriculture

The research team is now studying the processes regulated by MtWRKY69 to determine its mode of action. Phylogenetic analysis determined that the sRNA targeted sequence of MtWRKY69 is conserved through evolution, but specifically in those species capable of forming AM symbioses. Authors speculate that MtWRKY69 could be implicated in the plant’s stress response and that its downregulation could contribute to local suppression of the plant immune response, which would favor such an intimate root colonization by the beneficial fungus.

Improving the symbiotic capacity between plants and microorganisms offers a natural solution to many of the environmental problems we face. According to Ignacio Rubio-Somoza, ″This knowledge could lead to a ‘genomic Tinder’, matching plants and microorganisms to optimize symbiosis and address agricultural and environmental challenges.″

This discovery opens the door to potential applications in agriculture, including the reduction of the need for fertilizers and agrochemicals by enhancing natural nutrient uptake, or the development of crops with improved resilience to environmental stressors like drought, temperature fluctuations, pests, etc. In addition, with increasing evidence of mycorrhizal contributions to global carbon dynamics, enhancing plant-fungal symbioses offers a sustainable path forward in increasing CO2 fixation and mitigating climate crisis.

Reference article

Alessandro Silvestri*, William Conrad Ledford*, Valentina Fiorilli, Cristina Votta, Alessia Scerna, Jacopo Tucconi, Antonio Mocchetti, Gianluca Grasso, Raffaella Balestrini, Hailing Jin, Ignacio Rubio-Somoza and Luisa Lanfranco. A fungal sRNA silences a host plant transcription factor to promote arbuscular mycorrhizal symbiosis. New Phytologist, https://doi.org/10.1111/nph.20273