Large-scale transcriptomics to dissect two years of the life of a fungal phytopathogen interacting with its host plant

Abstract

The fungus Leptosphaeria maculans has an exceptionally long and complex relationship with its host plant, Brassica napus, during which it switches between different lifestyles, including asymptomatic, biotrophic, necrotrophic, and saprotrophic stages. The fungus is also exemplary of two-speed genome organisms in which gene-rich and repeat-rich regions alternate. Except for a few stages of plant infection under controlled conditions, nothing is known about the genes mobilized by the fungus throughout its life cycle, which may last several years in the field. We show here that about 9% of the genes of this fungus are highly expressed during its interactions with its host plant. These genes are distributed into eight well-defined expression clusters, corresponding to specific infection lifestyles or to tissue-specific genes. All expression clusters are enriched in effector genes, and one cluster is specific to the saprophytic lifestyle on plant residues. One cluster, including genes known to be involved in the first phase of asymptomatic fungal growth in leaves, is re-used at each asymptomatic growth stage, regardless of the type of organ infected. The expression of the genes of this cluster is repeatedly turned on and off during infection. Whatever their expression profile, the genes of these clusters are located in regions enriched in heterochromatin, either constitutive or facultative. These findings provide support for the hypothesis that fungal genes involved in niche adaptation are located in heterochromatic regions of the genome, conferring an extreme plasticity of expression. This work opens up new avenues for plant disease control, by identifying stage-specific effectors that could be used as targets for the identification of novel durable disease resistance genes, or for the in-depth analysis of chromatin remodeling during plant infection, which could be manipulated to interfere with the global expression of effector genes at crucial stages of plant infection.

A new family of structurally conserved fungal effectors displays epistatic interactions with plant resistance proteins

“…RNAseq data corresponding to different stages of infection of oilseed rape cotyledons by L. maculans as well as during axenic growth of L. maculans on V8 solid medium were recently generated (Gay et al, 2020), and expression kinetics of the LARS effector genes analyzed (Fig 4). All the genes showed over-expression during the biotrophic asymptomatic phase of cotyledon infection.…”

“…In L. maculans ‘brassicae’, thirteen LARS effectors could be detected and their expression during the primary biotrophic stages of oilseed rape cotyledon infection suggests they are bona fide effectors. They represent 14 % of the candidate effectors specifically overexpressed during the biotrophic stages of oilseed rape infection (nine LARS effectors among the 63 effector genes in Cluster 2 ‘biotrophy’ defined by Gay et al (2020)). The LARS family also comprises four out of the nine cloned AVR genes from L. maculans .…”

“…RNAseq data were previously generated by Gay et al (2020). The expression levels of structural family members were analyzed in L. maculans after extraction of RNAseq data corresponding to the following stages of infection: 2, 5, 7, 12 and 15 days after infection of oilseed rape cotyledons (susceptible cultivar Darmor-Bzh).…”

“…Expression kinetics of the LARS structural analogues identified in L. maculans 'brassicae'.Expression is based on RNAseq data described in[27] normalized by the total number of sequences per condition, counts per million (CPM), and represented on a logarithmic scale, following inoculation of cotyledons from a susceptible oilseed rape cultivar, Darmor-Bzh. Coty (cotyledons of the susceptible oilseed rape cultivar, Darmor-Bzh infected by L. maculans 'brassicae' 2, 5, 7, 9, 12 and 15 days after infection (DPI)); v8 (growth on solid medium V8 for one week).…”

A new avirulence gene of Leptosphaeria maculans, AvrLm14, identifies a resistance source in American broccoli (Brassica oleracea) genotypes

“…In relation to this genome location we also found recently that all AvrLm genes are enriched in H3K9me3 (trimethylation of lysine 9 of histone H3) marks, usually regarded as characteristic of constitutive heterochromatin in other organisms (Soyer et al, 2021). Being associated with this repressive mark, none of the AvrLm genes are expressed during vegetative growth (Soyer et al, 2021), but all of them show the same kinetics of expression when the fungus is inoculated onto cotyledons of B. napus, with a strong peak of overexpression 5-9 days postinoculation before symptoms are expressed (Figure 3) (Gay et al, 2021). Additionally, all of them encode small secreted proteins, often but not always enriched in cysteine residues, and showing no or few matches in databases, no recognizable domain, and no postulated function.…”