Phytophthora infestans Ago1-associated miRNA promotes potato late blight disease.

Phytophthora spp. cause serious damage to plants by exploiting a large number of effector proteins and small RNAs (sRNAs). Several reports have described modulation of host RNA biogenesis and defence gene expression. Here, we analysed Phytophthora infestans Argonaute (Ago) 1 associated small RNAs during potato leaf infection. Small RNAs were co-immunoprecipitated, deep sequenced and analysed against the P. infestans and potato genomes, followed by transcript analyses and transgenic assays on a predicted target. Extensive targeting of potato and pathogen-derived sRNAs to a range of mRNAs was observed, including 638 sequences coding for resistance (R) proteins in the host genome. The single miRNA encoded by P. infestans (miR8788) was found to target a potato alpha/beta hydrolase-type encoding gene (StABH1), a protein localized to the plasma membrane. Analyses of stable transgenic potato lines harbouring overexpressed StABH1 or artificial miRNA gene constructs demonstrated the importance of StABH1 during infection by P. infestans. miR8788 knock-down strains showed reduced growth on potato, and elevated StABH1 expression levels were observed when plants were inoculated with the two knock-down strains compared to the wild-type strain 88069. The findings of our study suggest that sRNA encoded by P. infestans can affect potato mRNA, thereby expanding our knowledge of the multifaceted strategies this species uses to facilitate infection.

A Verticillium longisporum pleiotropic drug transporter determines tolerance to the plant host ß-pinene monoterpene.

Terpenes constitute a major part of secondary metabolites secreted by plants in the rhizosphere. However, their specific functions in fungal-plant interactions have not been investigated thoroughly. In this study we investigated the role of monoterpenes in interactions between oilseed rape (Brassica napus) and the soilborne pathogen Verticillium longisporum. We identified seven monoterpenes produced by B. napus, and production of α-pinene, ß-pinene, 3-carene, and camphene was significantly increased upon fungal infection. Among them, ß-pinene was chosen for further analysis. Transcriptome analysis of V. longisporum on exposure to ß-pinene resulted in identification of two highly expressed pleotropic drug transporters paralog genes named VlAbcG1a and VlAbcG1b. Overexpression of VlAbcG1a in Saccharomyces cerevisiae increased tolerance to ß-pinene, while deletion of the VlAbcG1a homologous gene in Verticillium dahliae resulted in mutants with increased sensitivity to certain monoterpenes. Furthermore, the VlAbcG1a overexpression   strain displayed an increased tolerance to ß-pinene and increased virulence in tomato plants. Data from this study give new insights into the roles of terpenes in plant-fungal pathogen interactions and the mechanisms fungi deploy to cope with the toxicity of these secondary metabolites.

smartPARE: An R Package for Efficient Identification of True mRNA Cleavage Sites.

Degradome sequencing is commonly used to generate high-throughput information on mRNA cleavage sites mediated by small RNAs (sRNA). In our datasets of potato (Solanum tuberosum, St) and Phytophthora infestans (Pi), initial predictions generated high numbers of cleavage site predictions, which highlighted the need of improved analytic tools. Here, we present an R package based on a deep learning convolutional neural network (CNN) in a machine learning environment to optimize discrimination of false from true cleavage sites. When applying smartPARE to our datasets on potato during the infection process by the late blight pathogen, 7.3% of all cleavage windows represented true cleavages distributed on 214 sites in P. infestans and 444 sites in potato. The sRNA landscape of the two organisms is complex with uneven sRNA production and cleavage regions widespread in the two genomes. Multiple targets and several cases of complex regulatory cascades, particularly in potato, was revealed. We conclude that our new analytic approach is useful for anyone working on complex biological systems and with the interest of identifying cleavage sites particularly inferred by sRNA classes beyond miRNAs.