A structural homologue of the plant receptor D14 mediates responses to strigolactones in the fungal phytopathogen Cryphonectria parasitica.

Valentina Fiorilli,Giulia D’Arrigo,François‐Didier Boyer,David Cornu,Cristina Prandi,Marco Forgia,Luisa Lanfranco,Salim Al‐Babili,Massimo Turina,Francesca Spyrakis,Alexandre De Saint Germain,Francesca Cardinale,Philippe Le Bris

doi:10.1111/nph.18013

Abstract

Summary Strigolactones (SLs) are plant hormones and important signalling molecules required to promote arbuscular mycorrhizal (AM) symbiosis. While in plants an α/β‐hydrolase, DWARF14 (D14), was shown to act as a receptor that binds and cleaves SLs, the fungal receptor for SLs is unknown.Since AM fungi are currently not genetically tractable, in this study, we used the fungal pathogen Cryphonectria parasitica, for which gene deletion protocols exist, as a model, as we have previously shown that it responds to SLs. By means of computational, biochemical and genetic analyses, we identified a D14 structural homologue, CpD14.Molecular homology modelling and docking support the prediction that CpD14 interacts with and hydrolyses SLs. The recombinant CpD14 protein shows α/β hydrolytic activity in vitro against the SLs synthetic analogue GR24; its enzymatic activity requires an intact Ser/His/Asp catalytic triad. CpD14 expression in the d14‐1 loss‐of‐function Arabidopsis thaliana line did not rescue the plant mutant phenotype. However, gene inactivation by knockout homologous recombination reduced fungal sensitivity to SLs.These results indicate that CpD14 is involved in SLs responses in C. parasitica and strengthen the role of SLs as multifunctional molecules acting in plant–microbe interactions.

Highlights

Cryphonectria parasitica is a bark pathogen which causes perennial necrotic lesions on above-ground part of susceptible host trees
Gene inactivation by knock-out homologous recombination reduced fungal sensitivity to SLs. These results indicate that CpD14 is involved in SLs responses in C. parasitica and strengthen the role of SLs as multifunctional molecules acting in plant microbe-interactions
At 10-4 M the stereoisomers whose stereochemistry corresponds to natural canonical SLs, (+)-GR24 and ( )-2'-epiGR24 which have the D-ring with 2'R configuration, and the ( )- GR24 stereoisomer (2'S configuration), whose stereochemistry is not encountered in natural SLs, led to a reduction of fungal colony diameter at 72 and 96 h after inoculation (Supporting Information Fig. S1a)

Summary

Introduction

Cryphonectria parasitica is a bark pathogen which causes perennial necrotic lesions (cankers) on above-ground part of susceptible host trees This fungus is still a major threat to American and European chestnut trees causing the blight disease, which leads to yield losses of fruit and wood (Rigling & Prospero, 2018). A clear D14 homologue was not found in the genome of AM fungi (Tisserant et al, 2013; Lin et al, 2014), the most studied fungal system for SLs responses In this context, we decided to exploit the SLs-sensitive fungus C. parasitica for which, in contrast to AM fungi, stable genetic transformation protocols are available. We identified a candidate D14 structural homologue in C. parasitica and called it CpD14 (CpD14); by means of molecular modeling analyses and biochemical characterization of the recombinant protein we demonstrated CpD14 binding and enzymatic activity on natural SLs and their synthetic analogues. Gene inactivation by knock-out homologous recombination reduced C. parasitica sensitivity to SLs, indicating that the gene is involved in SLs responses in this fungus

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Discussion

Conclusion