Project description:Potato is one of the most important food crops for human consumption. The obligate biotrophic pathogen Spongospora subterranea infects potato roots and tubers, resulting in considerable loss of potato tuber yield and quality. A comprehensive understanding of how potato plants respond to S. subterranea infection is essential for the development of pathogen-resistant crops. Here we employed label-free proteomics and phosphoproteomics to quantify protein-level responses of the susceptible and resistant potato cultivars in response to S. subterranea. A total of 2669 proteins and 1498 phosphoproteins were quantified in the leaf samples of the different treatment groups. Following statistical analysis of the proteomic data, oxidoreductase activity, electron transfer, and photosynthesis were identified as significant processes that differentially changed upon infection specifically in the resistant cultivar and not in the susceptible cultivar. The phosphoproteomics results indicated increased activity of signal transduction and defence response functions in the resistant cultivar. In contrast, the majority of increased phosphoproteins in the susceptible cultivar were related to transporter activity and localisation. This study provides new insight into the molecular mechanisms involved in potato resistance to S. subterranea infection and has highlighted the critical roles of protein phosphorylation in the regulation of potato immune response.
Project description:We report the application of RNA- sequencing technology for high-throughput profiling of histone modifications in mammalian cellsor identification of expressed genes upon infection by Spongospora subterranea. Using RNA-sequencing (RNA-seq), 2058 differentially expressed genes (DEGs) were identified from two potato cultivars (tolerant and susceptible) in response to Sss infection. Analysis of the expression patterns of ten selected defense-response genes was carried out at two different stages of tuber growth using RT-qPCR to validate the RNA-seq data. Several defense related genes showed contrasting expression patterns between the tolerant and susceptible cultivars, including marker genes involved in the salicylic acid hormonal response pathway (StMRNA, StUDP and StWRKY6). Induction of six defense related genes (StWRKY6, StTOSB, StSN2, StLOX, StUDP and StSN1) persisted until harvest of the tubers, while three other genes (StNBS, StMRNA and StPRF) were highly up-regulated during the initial stages of disease development. The results of this study suggested that the tolerant potato cultivar employs quantitative resistance and salicylic acid pathway hormonal responses against tuber infection by Sss. The identified genes have the potential to be used in the development of molecular markers for selection of powdery scab resistant potato lines in marker assisted breeding programs.
Project description:This project is aiming to identify specific root surface proteins from susceptible and resistant potato strains and identify those factors responsible for Spongospora subterranea zoospore binding.
Project description:For potato crops, host resistance is currently the most effective and sustainable tool to manage potato root and tuber diseases caused by the plasmodiophorid, Spongospora subterranea. Arguably, zoospore root attachment is the most critical phase of the pathogen infection, however, the mechanisms underlying zoospore root attachment remains unknown. This study investigated the potential role of root cell wall surface polysaccharides and proteins in zoospore root attachment in resistant and susceptible potato cultivars. We first compared the effects of enzymatic removal of root cell wall proteins, N-linked glycans or polysaccharides on S. subterranea attachment to root tissue of resistant and susceptible potato cultivars. Subsequently, mass spectrometry analysis of peptides released by trypsin shaving (TS) of root segments identified 1235 proteins, of which 262 were differentially abundant between the resistant and susceptible cultivars. In particular, proteins associated with glutathione metabolism and lignin biosynthesis were more abundant in the resistant cultivar. Comparison with whole-root proteomic analysis of the same resistant and susceptible cultivars led to identification of 226 proteins unique to the TS dataset, of which 188 were significantly different between cultivars. Among these, the pathogen defence-related cell wall protein stem 28 kDa glycoprotein and two major latex proteins were significantly less abundant in the resistant cultivar compared to the susceptible cultivar. A further major latex protein was detected at reduced levels in the resistant cultivar in both TS and whole-root proteomic datasets. In contrast, in the TS-specific dataset, three glutathione S-transferase proteins were more abundant in the resistant cultivar, while the protein glucan endo-1,3-beta-glucosidase was significantly increased in both the TS and whole-root datasets. These results imply a particular role of major latex proteins and glucan endo-1,3-beta-glucosidase in the regulation of host susceptibility to S. subterranea.
Project description:The potato powdery scab agent Spongospora subterranea causes damage on the skin of tubers and induces root gall formation, precipitating considerable yield and quality losses. Currently, there are no effective chemical treatments for the control of powdery scabs. Understanding the inducible defence responses in roots of potato plants in the resistant and susceptible host environment, particularly during colonisation of the root by S. subterranea is required for the breeding of novel resistant cultivars. Here, we integrated transcriptomics, proteomics and metabolomics datasets to uncover the mechanisms underlying of the potato resistance to powdery scab. This multi-omics approach identified upregulation of glutathione metabolism at the levels of RNA, protein and metabolite in the resistant cultivar but not in the susceptible cultivar. Upregulation of the lignin metabolic process was also specific to in the resistant cultivar at the transcriptome level. In addition, Tthe inositol phosphate pathway was differentially expressed between two cultivars in response to S. subterranea infection, where it was upregulated in the susceptible cultivar but downregulated in the resistant cultivar. We provide, for the first time, large-scale multi-omics data of Spongospora-potato interaction, thereby suggesting the signaling role of glutathione metabolism in the potato resistance against powdery scab
Project description:Spongospora subterranea is an obligate biotrophic pathogen, causing a tremendous economic loss in the potato industry. Currently, there are no effective chemical or biological strategies for the control of S. subterranea. Understanding the gene regulation of pathogens in their host is dependent on multidimensional datasets. To further our understanding of S. subterranea biology during infection, we characterized the transcriptome and proteome of the pathogen inside the susceptible and resistant potato cultivars. A total of 7650 transcripts from S. subterranea were identified in the transcriptome analysis in which 1377 transcripts were differentially expressed between two cultivars. In proteome analysis, we identified 117 proteins with 14 proteins significantly changed in comparison between resistant and susceptible cultivars. The transcriptome analysis uncovered the gene regulatory modules underlying virulence. The functional annotation of transcriptome data indicated that the gene ontology terms related to the transportation and actin processes were induced in the resistant cultivar. The downregulation of enzyme activity and nucleic acid metabolism in the resistant cultivars suggesting a remarkable influence of these processes in the virulence of S. subterranea. The protein analysis results indicated that the majority of identified proteins were related to the metabolic processes. The present study provides a comprehensive molecular insight into the multiple layers of gene regulation that contribute to S. subterranea germination and growth in planta and illuminates the role of host immunity in affecting pathogen responses.
Project description:Spongospora subterranea is a soil-borne plant pathogen responsible for economically significant root and powdery scab diseases of potato. However, Tthe obligate biotrophic nature of S. subterranea has made detailed study of the pathogen problematic. Here, we first compared the benefits of sporosori partial purification utilizing Ludox gradient centrifugation. We then undertook optimization efforts for protein isolation comparing the use of a urea buffer followed by Single-Pot Solid-Phase-enhanced Sample Preparation (SP3) and a sodium dodecyl sulfate (SDS) buffer followed by suspension-trapping (S-Trap). Label-free, quantitative proteomics was then used to evaluate the efficiency of sporosori purification and protein preparation methods. The purification protocol produced a highly purified suspension of S. subterranea sporosori without affecting the viability of the spores. Results indicated that the use of a combination of SDS and S-Trap for sample clean-up and digestion obtained a significantly higher number of identified proteins compare to using urea and SP3, with 218 and 652 proteins identified using SP3 and S-Trap methods, respectively. Analysis of proteins by mass spectrometry showed that the number of identified proteins increased by approximately 40% after the purification of spores by Ludox. These results suggested a potential use of the described spore purification and protein preparation methods for proteomics study of obligate biotrophic pathogens such as S. subterranea.
Project description:Identification of differentially expressed genes in tolerant and susceptible potato cultivars in response to Spongospora subterranea f. sp. subterranea tuber infection