Bioinformatic and functional characterisation of Globodera pallida effector genes

Pathogens secrete molecules, termed effectors, to manipulate their host to the benefit of the pathogen. Effectors of plant parasitic nematodes are predicted to have a range of functions such as facilitating invasion, initiation and maintenance of the feeding site, and suppression of host defences. The genome sequence of the potato cyst nematode Globodera pallida was analysed to identify putative effectors. They include: 129 effectors similar to those previously identified from cyst nematodes, 53 cell wall modifying enzymes and 117 novel putative effectors. Only four effectors were common between G. pallida and the root-knot nematode Meloidogyne incognita. These could have a conserved role in plant parasitism. A large SPRY domain containing gene family was identified in G. pallida. It has 299 members, of which 30 are predicted to be secreted and therefore categorised as effectors. Phylogenetic analysis showed that the family is hugely expanded and specific to Globodera species. Fifty-four putative effectors were cloned from G. pallida cDNA. Transgenic lines of Arabidopsis thaliana and Solanum tuberosum L. ‘Désirée’ were produced, to express a range of these effectors and act as tools for functional characterisation. Potato lines that expressed selected effectors were subjected to phenotypic analysis and pathogen susceptibility assays. The largest range of aberrant phenotypes was observed for those plants expressing GpIA7 and GpIVG9. Potato lines expressing GpIA7 showed altered growth phenotypes and an increased susceptibility to Phytophthora infestans CS-12. GpIVG9-expressing potato lines showed accelerated growth, distorted leaves and increased susceptibility to nematode invasion. A more in-depth functional characterisation was conducted on a ubiquitin extension protein effector. The G. pallida ubiquitin extension protein suppressed PAMPtriggered immunity and the C-terminal extension was required for this activity. The outcomes from this work and the tools generated for future experimentation will contribute to elucidating the complex interactions between pathogens and their hosts.