Plasmodiophora brassicae causes clubroot disease in cruciferous. In this report, lipid droplets were observed in the resting spores of P. brassicae. 295 lipid droplet-associated proteins were identified and categorized into nine groups. Transcriptome analysis of these proteins during three different zoosporic stages revealed differences in gene expression pattern. GO enrichment analysis revealed that these proteins associated with lipid droplets were mainly linked to biosynthesis and metabolism. GC-MS analysis revealed that lipid droplets contain seven types of free fatty acids: saturated fatty acids C16:0 and C18:0, and unsaturated fatty acids C18:1Δ9, C18:1Δ11, C18:2, C20:4 and C20:5. P. brassicae accumulated a large amount of triacylglycerols (TAGs). We systematically analyzed the putative proteins involved in TAG biosynthesis and its metabolic pathway. KEGG pathway analysis defined 3390 genes, including 167 genes involved in lipid metabolism. Transcriptome analysis revealed that 162 candidate enzymes involved in lipid metabolism were differential expressed. Our omics studies are the first to investigate the lipid droplet organelles in P. brassicae, providing a reference resource to study protist lipid droplets.

Botrytis-induced kinase1 (BIK1), a receptor-like cytoplasmic kinase, plays an important role in resistance against pathogens and insects in Arabidopsis thaliana. However, it remains unknown whether BIK1 functions against Plasmodiophora brassicae, an obligate biotrophic protist that attacks cruciferous plants and induces gall formation on roots. Here, we investigated the potential roles of receptors FLS2, BAK1, and BIK1 in the infection of P. brassicae cruciferous plants. Wild-type plants, fls2, and bak1 mutants showed typical symptom on roots, and the galls were filled with large quantities of resting spores, while bik1 mutant plants exhibited strong resistance to P. brassicae. Compared with that of the wild-type plants, the root hair and cortical infection rate of bik1 mutant were significantly reduced by about 40–50%. A considerable portion of bik1 roots failed to form typical galls. Even if some small galls were formed, they were filled with multinucleate secondary plasmodia. The bik1 plants accumulated less reactive oxygen species (ROS) at infected roots than other mutants and wild-type plants. Exogenous salicylic acid (SA) treatment alleviated the clubroot symptoms in wild-type plants, and the expression of the SA signaling marker gene PR1 was significantly increased in bik1. Both sid2 (salicylic acid induction-deficient 2) and npr1-1 [non-expresser of PR genes that regulate systemic acquired resistance (SAR)] mutants showed increased susceptibility to P. brassicae compared with wild-type plants. These results suggest that the resistance of bik1 to P. brassicae is possibly mediated by SA inducible mechanisms.

Plasmodiophora brassicae is an important biotrophic eukaryotic plant pathogen and a member of the rhizarian protists. This biotrophic pathogen causes clubroot in cruciferous plants via novel intracellular mechanisms that are markedly different from those of other biotrophic organisms. To date, genomes from six single spore isolates of P. brassicae have been sequenced. An accurate description of the evolutionary status of this biotrophic protist, however, remains lacking. Here, we determined the draft genome of the P. brassicae ZJ-1 strain. A total of 10,951 protein-coding genes were identified from a 24.1 Mb genome sequence. We applied a comparative genomics approach to prove the Rhizaria supergroup is an independent branch in the eukaryotic evolutionary tree. We also found that the GPCR signaling pathway, the versatile signal transduction to multiple intracellular signaling cascades in response to extracellular signals in eukaryotes, is significantly enriched in P. brassicae-expanded and P. brassicae-specific gene sets. Additionally, treatment with a GPCR inhibitor relieved the symptoms of clubroot and significantly suppressed the development of plasmodia. Our findings suggest that GPCR signal transduction pathways play important roles in the growth, development, and pathogenicity of P. brassicae.

BackgroundThe eukaryotic unicellular protist Plasmodiophora brassicae is an endocellular parasite of cruciferous plants. In host cortical cells, this protist develops a unicellular structure that is termed the plasmodium. The plasmodium is actually a multinucleated cell, which subsequently splits and forms resting spores. The mechanism for the growth of this endocellular parasite in host cell is unclear.ResultsHere, combining de novo genome sequence and transcriptome analysis of strain ZJ-1, we identified top five significant enriched KEGG pathways of differentially expressed genes (DEGs), namely translation, cell growth and death, cell communication, cell motility and cancers. We detected 171 proto-oncogenes from the genome of P. brassicae that were implicated in cancer-related pathways, of which 46 were differential expression genes. Three predicted proto-oncogenes (Pb-Raf1, Pb-Raf2, and Pb-MYB), which showed homology to the human proto-oncogenes Raf and MYB, were specifically activated during the plasmodial growth in host cortical cells, demonstrating their involvement in the multinucleate development stage of the unicellular protist organism. Gene networks involved in the tumorigenic-related signaling transduction pathways and the activation of 12 core genes were identified. Inhibition of phosphoinositol-3-kinase relieved the clubroot symptom and significantly suppressed the development process of plasmodia.ConclusionsProto-oncogene-related regulatory mechanisms play an important role in the plasmodial growth of P. brassicae.Electronic supplementary materialThe online version of this article (10.1186/s12864-018-5307-4) contains supplementary material, which is available to authorized users.

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