If plant-plant interactions and allorecognition mechanisms are interesting to you, you should apply to our PhD position at the GAIA grad school, Institut Agro Montpellier! How plants alter immunity in their neighbours?
PhD project: How do plants modify disease resistance in their neighbors? Omics approaches for the identification of genes and molecules involved in the regulation of immunity by plant-plant interactions
Keywords: Plant-Plant interactions, Wheat, Leaf rust, Comparative transcriptomic, GWAST. hemes Domain: Mechanisms of plant-plant interactions in the context of pathogen infection. Plant pathology
Profile and skills required:
Candidates must have a master degree (or equivalent) in plant physiology including notions of plant pathology. Knowledge in genomics and molecular biology is highly desirable, as well as a good mastery of statistics in R.
The deleterious impact of agricultural systems resulting from the Green Revolution, based on the use of phytosanitary products and a small number of elite varieties, has contributed significantly to the loss of biomass and biodiversity in ecosystems (Degiorgi and Badot, 2020). Several strategies exist to remedy the problems associated with this type of agriculture, including the use of varietal mixtures.
Indeed, mixed crops require less nitrogen inputs and phytosanitary products than pure crops while at the same time promoting a stabilization and/or a slight increase in yields, especially in the wild ancestors of cultivated species (Chacón-Labella et al., 2019). In agricultural plots, the positive impact of varietal mixtures on epidemic control or resistance maintenance has been widely demonstrated (Kristoffersen et al., 2020; Montazeaud et al., 2022; Wuest et al., 2021). However, this general benefit is not systematic and can be suppressed by specific allele combinations (Montazeaud et al., 2022), highlighting the need to know the genetic basis of neighbor-regulated immunity to better use mixtures.
Recent results from the MOMIE team at the Plant Health Institute of Montpellier have shown that certain wheat varieties modulate the susceptibility of other wheat varieties against leaf rust. According to these initial results, neighbor-regulated plant immunity relies on inter-root communication events that are active even in the absence of an epidemic (Pélissier, Buendia, et al., 2021; Pélissier, Violle, et al., 2021). One of our hypotheses is that this inter-root communication would be ensured by specialized metabolites exuded in the rhizosphere that have been described in the context of allelopathic mechanisms (Li et al., 2020; Wheeldon et al., 2022; Yoneyama et al., 2022) but whose action on the regulation of immunity has not yet been demonstrated.
Other phenomena could be involved, such as nutrient competition in the root compartment, but the molecular mechanisms by which immunity is regulated under mixed conditions remain to be described. This project will use wheat as a study model, especially because its specialized metabolism is poorly characterized, but presents intriguing specificities regarding the nature of the biosynthesized molecules, especially in response to pathogens (Polturak et al., 2022).
Bread wheat also has a high quality assembled genome, as well as a large resource of gene expression data (>1,000 transcriptomes) (Ramírez-González et al., 2018; Walkowiak et al., 2020) and mutant library facilitating functional validations (Krasileva et al., 2017). In addition, mixtures of varieties are growing rapidly in France in wheat since they already represent 15-20% of the area (~1 million hectares). We will use leaf rust as pathogen for the ease of quantification of symptoms and because this pathogen is responsible for a loss of several billion worldwide (Beddow et al., 2015; Huerta-Espino et al., 2011).
Through comparative transcriptomic approaches (primary objective), association genetics (GWAS, secondary objective), and analysis of root exudoma (ternary objective), this thesis project aims to identify genes and molecules involved in the regulation of neighborhood immunity in soft wheat. For comparative transcriptomics, the focal soft wheat variety Cadenza will be used because a homozygous point mutant library is available (Krasileva et al., 2017). Cadenza will be co-cultivated with itself (pure condition or control) or with each of the other varieties of a panel of 200 soft wheat accessions including traditional varieties with little selection “landrace”, old and recent cultivars, as well as currently cultivated elite varieties.
This panel of varieties has been used to trace the history of soft wheat domestication for 10,000 years (Pont et al., 2019). After inoculation of foliage with a strain of brown rust, the percentage of leaf area showing symptoms will be quantified for the 201 conditions, in triplicate. The quantitative and statistical analysis of the symptoms in mixed conditions compared to pure will allow the identification of 20 neighboring varieties positively or negatively affecting the susceptibility of Cadenza to brown rust. Based on this phenotyping, the
Cadenza transcriptomes will be analyzed in the presence of 30 different culture environments. In the presence of 10 varieties reducing its susceptibility to rust, in the presence of 10 varieties having no influence on its susceptibility (including the pure condition) as well as in the presence of 10 varieties increasing its susceptibility. Clusters of co-expression in these three conditions will be obtained, thus providing lists of transcripts encoding proteins potentially involved in the immune response of the focal plant to its neighborhood. The comparison of these data with the public data of gene ontology and transcriptomics of common wheat will allow the establishment of a shortlist of candidate genes. These genes
Candidates will be functionally tested by reverse genetics using Cadenza’s point mutant library. This first part constitutes the main objective of the thesis. At the same time, an association genetic analysis of the 200 neighboring accessions (Pélissier et al., 2023) could make it possible to identify genomic intervals containing SNPs associated with the immune phenotype. The haplotypes of these QTLs will be identified and validated for their effect on the susceptibility of Cadenza by co-cultivating it with ad hoc varieties possessing these haplotypes. To identify focal transcripts whose translation is modified by a validated neighboring haplotype, an analysis of the reductome and the Cadenza transcriptome in co-culture with the validated haplotypes will be undertaken.
Genes whose translation is regulated will be functionally tested by driving Cadenza point mutants. Under these same mixing conditions, an analysis of the root transcriptome of the haplotypes and the focal will be carried out to identify the most active metabolic pathways in order to propose candidate metabolites. The GWAS approach described above constitutes the second main objective of the thesis, and all the proposals below will be optional depending on the progress and interest of the recruited doctoral student. Recently, positional cloning being long and tedious for a polyploid species, a strategy of gene silencing induced by viral RNA (VIGS) could be undertaken to identify the causative gene(s) within the QTLs most explanatory of susceptibility. of the focal plant.
Third, the development of an exploratory approach could allow the doctoral student to obtain very innovative results. A metabolomic and proteomic analysis of the roots or root exudates of the same couples as those used in transcriptomics will be carried out for the identification of signals involved in plant-plant interactions. To this end, the plants will be grown in hydroponics, for easy access to roots and exudates (Ghatak et al., 2022).
More specifically, pools of exudates or roots corresponding to the transcriptomic samples will be analyzed for their metabolite and protein content by untargeted mass spectrometry. The differential analysis with the neutral condition as a control will lead to the quantitative and statistical identification of molecules and proteins potentially involved in an inter-root communication involved in a modulation of immunity. The candidate molecules could be purified by preparative liquid chromatography and tested by exogenous treatment for their effect on the resistance of wheat to rust.
Candidate peptides will be synthesized and tested in the same way. This part of the thesis project will need optimization at several levels (culture conditions, preparation and purification of exudates, non-targeted metabolomics and identification of compounds), and will therefore be a support project for comparative transcriptomics and GWAS whose necessary tools are mastered in the MOMIE team.
Applications deadline: 28 April 2023