Molecular investigation of Rlm3 from rapeseed as a broad-spectrum resistance gene against fungal pathogens producing structurally conserved effectors

Recognition of a pathogen avirulence (AVR) effector protein by its cognate plant resistance (R) protein triggers immune responses that render the plant resistant, representing an efficient disease control strategy. While (AVR) effectors have long been considered species- or genotype-specific, based on a lack of homology in sequence databases, a growing number of studies have now shown that these proteins belong to a limited set of structural families. This is an important finding because it paves the way for the identification or engineering of broad- spectrum R proteins capable of recognizing several members of the same structural family. In the Leptosphaeria maculans / rapeseed ( Brassica napus ) pathosystem, 13 AVR genes have been cloned, of which four encode effectors belonging to the LARS ( Leptosphaeria AviRulence and Supressing) structural family that has also been found in thirteen other phytopathogenic fungi. Homologues of the L. maculans AvrLm3 AVR protein, a LARS family member, have been identified in other fungal species, including an AVR protein from Fulvia fulva called Ecp11-1. We have previously shown that Ecp11-1 is recognized by rapeseed varieties carrying the Rlm3 R gene, and that this recognition is masked by the presence of another LARS AVR gene, AvrLm4-7 . In this study, we expanded our characterization of the Rlm3 resistance spectrum to putative effectors from Fusarium oxysporum and Zymoseptoria ardabiliae and showed that one effector from F. oxysporum f. sp. narcissi behaves like Ecp11-1, being recognized by Rlm3, and this recognition also being masked by the presence of AvrLm4-7 . We finally investigated which protein regions and amino acids were necessary for the recognition of AvrLm3 and Ecp11-1 by Rlm3. This analysis is a first step towards the identification or engineering of broad- spectrum R proteins that confer protection against multiple phytopathogens through the recognition of structural effector families.