Metabolomic approaches reveal that cell wall modifications play a major role in ethylene-mediated resistance against Botrytis cinerea

In Arabidopsis, resistance to the necrotrophic fungus Botrytis cinerea is conferred by ethylene via poorly understood mechanisms. Metabolomic approaches compared the responses of the wild-type, the ethylene-insensitive mutant etr1-1, which showed increased susceptibility, and the constitutively active ethylene mutants ctr1-1 and eto2 both exhibited decreased susceptibility to B. cinerea. Fourier transform-infrared (FT-IR) spectroscopy demonstrated reproducible biochemical differences between treatments and genotypes. To identify discriminatory mass-to-charge ratios (m/z) associated with resistance, discriminant function analysis was employed on spectra derived from direct injection electrospray ionisation-mass spectrometry on the derived principal components of these data. Ethylene-modulated m/z were mapped onto Arabidopsis biochemical pathways and many were associated with hydroxycinnamate and monolignol biosynthesis, both linked to cell wall modification. A high-resolution linear triple quadrupole-Orbitrap hybrid system confirmed the identity of key metabolites in these pathways. The contribution of these pathways to defence against B. cinerea was validated through the use of multiple Arabidopsis mutants. The FT-IR microspectroscopy indicated that spatial accumulation of hydroxycinnamates and monolignols at the cell wall to confine disease was linked ot ethylene. These data demonstrate the power of metabolomic approaches in elucidating novel biological phenomena, especially when coupled to validation steps exploiting relevant mutant genotypes.