Core effectors and host targets of plant parasitic nematodes

Plant-parasitic nematodes (PPN) cause damage to all major crops across the world. The most damaging PPN are the sedentary endoparasitic species. One of the major clades of PPN includes the cyst nematodes (Globodera/Heterodera spp.), the reniform nematode Rotylenchulus reniformis and the false root-knot nematode Nacobbus aberrans. These nematodes induce a large multinucleate, metabolically active feeding site in the root of their host plants. Although different in their morphology, each is a type of syncytium, formed by the breakdown of the cell wall between the initial cell selected by the nematode and its neighbours followed by fusion of the protoplasts. The nematodes can only induce one syncytium in their life cycle. The syncytium must therefore be kept alive and protected from host defence responses in order to provide the nutrients required by the nematode. The interactions of PPN with their hosts, including the induction and upkeep of the feeding site, are mediated by effectors; proteins secreted by the nematode that manipulate host biochemistry to benefit the parasite. Effectors have been identified in a range of PPNs and are often present as large gene families. Understanding the functional roles of effectors and the molecular basis by which syncytia are induced/protected offers the prospects of new control strategies against these parasites. The supervisors of this project have been involved in the generation of genome/transcriptome data from Globodera pallida and G. rostochiensis, R. reniformis and N. aberrans. We have developed bioinformatic pipelines allowing for candidate effectors to be identified from these data. The main aim of this project is to use the data to allow the identification of "core" host proteins or processes, manipulated by all species, which by their very nature are likely to be key players in the interaction between the nematode and its host plant. To achieve this, the targets of "core" effectors, conserved between all three nematode groups, will be identified. The main work areas that are foreseen for the early part of the project are as follows; Identification of "core" effectors: The three genera identified span two key bifurcations in the phylum, and have large sequence databases, including genome and transcriptome information. All genes from the three nematode groups will be clustered, and those containing sequences from all groups which contain previously identified effectors will be highlighted. In addition, putative novel conserved effector clusters will be identified by the presence of signatures used to distinguish effectors from house-keeping genes (signal peptide for secretion and transcriptional profile consistent with a role in parasitism). In both cases, these analyses will be validated by in situ hybridisation to confirm that these sequences are expressed in nematode tissues with the capacity to secrete proteins into plants. Identification of conserved host targets/processes: All three genera can infect some common hosts, e.g. tomato and potato, and produce feeding sites with similar features. It is therefore likely that common host proteins or process must be manipulated. Initially focusing on those conserved effectors between species maximises the likelihood of identifying such 'key players'. Two complementary strategies will be used. Firstly, tagged effectors over expressed in planta will be pulled down, and any accompanying host proteins bound to the effectors will be identified by mass spectrometry. Secondly, yeast-2-hybrid libraries (potato and tomato) will be screened with effectors. Interactions will be confirmed by co-immunoprecipitation and/or FRET analysis. The later stages of the work will depend on the data obtained in the first phases. Key experiments may include functional analysis of the roles of conserved targets in nematode infection through over-expression, silencing, or genome editing. They may also include structural analysis of effector-target interactions.