Genome‐wide transcriptomic analyses provide insights into the lifestyle transition and effector repertoire of Leptosphaeria maculans during the colonization of Brassica napus seedlings

Abstract

Molecular interaction between the causal agent of blackleg disease, Leptosphaeria maculans (Lm), and its host, Brassica napus, is largely unknown. We applied a deep RNA-sequencing approach to gain insight into the pathogenicity mechanisms of Lm and the defence response of B. napus. RNA from the infected susceptible B. napus cultivar Topas DH16516, sampled at 2-day intervals (0-8 days), was sequenced and used for gene expression profiling. Patterns of gene expression regulation in B. napus showed multifaceted defence responses evident by the differential expression of genes encoding the pattern recognition receptor CERK1 (chitin elicitor receptor kinase 1), receptor like proteins and WRKY transcription factors. The up-regulation of genes related to salicylic acid and jasmonic acid at the initial and late stages of infection, respectively, provided evidence for the biotrophic and necrotrophic life stages of Lm during the infection of B. napus cotyledons. Lm transition from biotrophy to necrotropy was also supported by the expression function of Lm necrosis and ethylene-inducing (Nep-1)-like peptide. Genes encoding polyketide synthases and non-ribosomal peptide synthetases, with potential roles in pathogenicity, were up-regulated at 6-8 days after inoculation. Among other plant defence-related genes differentially regulated in response to Lm infection were genes involved in the reinforcement of the cell wall and the production of glucosinolates. Dual RNA-sequencing allowed us to define the Lm candidate effectors expressed during the infection of B. napus. Several candidate effectors suppressed Bax-induced cell death when transiently expressed in Nicotiana benthamaina leaves.

Comparative Transcriptomic Analysis of Virulence Factors in Leptosphaeria maculans during Compatible and Incompatible Interactions with Canola

“…On the basis of visual observations, this condition was clearly met as Topas-wild plants exhibited clear symptoms of infection over the course of the experiment that culminated with the presence of extensive necrotic tissues at 11 dpi while the infection never extended beyond the point of inoculation in Topas -Rlm2 plants. Other studies have reported similar symptom progression at varying time points (Lowe et al, 2014; Haddadi et al, 2016), thus suggesting that experimental conditions can influence the rapidity with which L. maculans can infect its host. For this reason, it is critical to ensure that sampling procedures include enough replications that will capture an accurate biological variability within a condition.…”

“…dpi, days post inoculation; FPKM, Fragments per kilo-base of transcript per million mapped reads; 1, (Haddadi et al, 2016); 2, (Lowe et al, 2014) .…”

“…The L. maculans -canola compatible interaction has been previously addressed using the RNA-seq approach (Lowe et al, 2014; Haddadi et al, 2016). These studies made significant efforts toward the understanding of susceptible reaction in canola after L. maculans infection and provides the first in-depth look into the transcriptomic profile of this interaction.…”

Impact of biotic and abiotic factors on the expression of fungal effector-encoding genes in axenic growth conditions

“…Despite its up-regulation in our in vitro experiment, Lema086540 was not retained as its log 2 -fold change was of only 1.8 and thus below our threshold. Two other genes (Lema006106, Lema084480), recently found to encode for SSP required for full or partial suppression of cell death (Haddadi et al, 2016) and located in GC-isochores were also up-regulated in presence of cycloheximide (Supplemental Table S2). In contrast, other avirulence genes located in AT-isochores such as AvrLm6 and AvrLm2 were not significantly affected by the cycloheximide treatment (data not shown).…”

Host–pathogen interactions in relation to management of light leaf spot disease (caused by Pyrenopeziza brassicae) on Brassica species

“…Transcriptome sequencing has been used to analyse the interaction between B. napus and L. maculans (Lowe et al 2014;Haddadi et al 2016); both studies have used susceptible cultivars to determine pathogen and host gene expression. Such studies are useful to determine potential pathogenicity (e.g.…”