Award details

Evolution of D14L signalling specificity for symbiosis and development

Reference	BB/V004263/1
Principal Investigator / Supervisor	Dr Emma Wallington
Co-Investigators / Co-Supervisors
Institution	National Inst of Agricultural Botany
Department	Centre for Research
Funding type	Research
Value (£)	114,999
Status	Current
Type	Research Grant
Start date	01/10/2021
End date	30/09/2024
Duration	36 months

Abstract

Rice offers a highly attractive plant system for an integrated combination of molecular, genetic and transcriptomic approaches, having a well annotated genome, efficient transformation protocols and its roots being readily colonised by AM fungi. With the proposal aiming at the elucidation of D14L protein characteristics driving signalling specificities, the technical workplan largely builds on metagenomics, plant transformation and phenotyping, including microscopic, biochemical and molecular phenotyping. The use of the fast and sensitive protein aligner DIAMOND2 enables another dimension of phylogenomics studies, due to the significantly accelerated analysis of the ever increasing, sequence data. In combination a newly developed algorithms to define protein similarity networks, the dynamics of evolvability of proteins, protein domains, or protein residues is revealed, and will then be overlaid with the taxonomic map of plant symbiotic competence. Computational followed by genetic validation reveals D14L sequence features that are significantly associated with the symbiosis signalling phenotype. Transformation of cereal crops is efficiently running in the team of the joint-applicant Emma Wallington (EW) who routinely delivers a high number of transformants with, in rice, at least 40% of lines carrying single copy T-DNA integrations. We base all our constructs for genetic complementation on synthesising and cloning the coding region of the D14L variants under the transcriptional control of the same native rice D14L promoter and terminator. Central phenotyping methodologies are rice germination to measure mesocotyl length, conventional light microscopy for the quantification of root colonisation, standard protoplast transformation and Western blotting to monitor protein abundance, RNAseq (Novogene) to produce the quantitative and qualitative estimate of transcripts.

Summary

A key event in the evolution of complex life on earth is the transition of plants from water to land. To conquer the terrestrial environment plants had to develop strategies for the acquisition of soil nutrients with initially primitive root precursors. Ever since, plants have lived in symbioses with nutrient-delivering beneficial fungi. Today, the arbuscular mycorrhizal (AM) symbiosis is the most commonly occurring beneficial plant-fungal interaction on Earth, contributing to plant fitness, plant biodiversity and global nutrient cycles. Yet, we are only beginning to unveil the molecular processes leading to the establishment of this intimate association and to control it for modern crop breeding. The Paszkowski group has found that an alpha/beta hydrolase receptor, called DWARF 14 LIKE (D14L), is essential for fungal perception of rice in the rhizosphere. The activation of this receptor is necessary for the unblocking of genetic programmes that enable symbiosis development and induce production of the phytohormone strigolactone (Gutjahr et al., Science 2015; Choi et al., Nat Commun 2020). The receptor is present across the plant kingdom and serves multifunctional roles, including the detection of the smoke constituent karrikin and the modulation of seedling development amongst others. However, D14L receptors from different plant species are functionally not necessarily equivalent. The Paszkowski lab recently found that the D14L homologue from Arabidopsis thaliana was only able to complement the developmental phenotype of rice d14l mutants, but failed to restore AM symbiosis. As D14L receptors are present across terrestrial plants, including early divergent plants and even Charophyte algae, they may have distinctly and successively diversified to acquire the signalling roles we observe today. We now wish to reproduce the evolutionary trajectory in order to define the protein features that condition signalling specificity, and to better understand their emergence.Exploiting the exponentially growing genome sequence resource, we propose here a high-resolution metagenomics approach, employing and developing computational tools that are significantly superior relative to conventional practices. This includes the development of new algorithmic steps enabling 10,000-fold computational acceleration compared to previous approaches. Results obtained from these in silico efforts are then functionally validated in rice to confirm their relevance in vivo. Insights generated here are vital for understanding how receptor functioning integrates and specifies different signalling cues. In doing so, this will open up new areas of research in computational and plant biology and provide a paradigm for which other examples of signalling processes can be assessed.

Committee	Research Committee B (Plants, microbes, food & sustainability)
Research Topics	Crop Science, Microbiology, Plant Science, Systems Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

Associated awards:

BB/V006029/1 Evolution of receptor signalling specificity for symbiosis and development

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