Interplay of dirgent and jacalin-proteins in broad-spectrum disease resistance of plants against fungal pathogens

• Applicants: Dr. Thomas Classen; Professor Dr. Ulrich Schaffrath
• Subject Area: Organismic Interactions, Chemical Ecology and Microbiomes of Plant Systems ; Plant Biochemistry and Biophysics ; Plant Physiology ; Plant Breeding and Plant Pathology
• Term: from 2017 to 2022
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 369034981

Fusion proteins consisting of a jacalin-related lectin (JRL) and a dirigent domain occur exclusively within the plant family Poaceae. The group of Ulrich Schaffrath has shown that constitutive over-expression of different members of this family in rice, barley and wheat, respectively, led to quantitative broad-spectrum disease resistance against different pathogens including bacteria, oomycetes and fungi. Recombinant proteins with an artificially separated JRL-domain localizes towards the site of penetration when expressed in barley attacked by the powdery mildew fungus. When co-expressed, also a protein consisting of the separated dirigent domain localizes at the penetration site together with the JRL-protein. The activity of both protein partners is required for enhanced pathogen resistance.Based on these results we hypothesize, that JRL- and dirigent proteins are part of an ancient plant defense system which consists of two components, one acting on pathogen recognition and the other on execution of resistance. Fusion of the respective proteins is an invention solely realized in Poaceae species. In the proposed project we will challenge this hypothesis by answering the following questions: i) which carbohydrate signature is exposed during pathogen attack and binds the JRL-domain; ii) which antimicrobial substance is produced in the presence of the dirigent-domain protein; and iii) are there pairs of JRL and dirigent proteins in other plant species that cooperatively act in the proposed manner. In particular, domain swap experiments will be performed with the domains of different Poaceae fusion proteins to unravel functional similarities. The identification of the JRL-domain binding partner and characterization of the biochemical reaction in which the dirigent-domain is involved will be targeted by the group of Thomas Classen, a biochemist. This will include synthesis of recombinant proteins, crystallography, pull-down assays of JRL-lectins with carbohydrates from pathogens and/or plants as well as HPLC-analyses.Arabidopsis will be used as model plant, to search for pairs of proteins that act in disease resistance in a similar manner as the above mentioned Poaceae-specific fusion proteins. Using bioinformatics, we already identified two promising candidate genes which are located next to each other on Arabidopsis chromosome 1. Both proteins will be expressed in barley to elucidate their involvement in penetration resistance. Promising candidates of JRL- and dirigent proteins will be used to construct an artificial fusion protein and test whether this is capable to provide broad-spectrum disease resistance. This finally will prove if translational approaches, in which artificial fusion proteins are expressed in crop plants, are a promising strategy to improve plant stress resistance.The combined expertise of the Schaffrath and Classen labs in molecular phytopathology and biochemistry will build a solid base for the realization of the project.

• DFG Programme: Research Grants