Deploying virus resistance in brassicas and understanding the interaction between the viral VPg protein and brassica eIF4E and eIF(iso)4E proteins.

It has been shown that the isoform of the eukaryotic translation initiation factor 4E (eIF(iso)4E) protein of Arabidopsis binds with the viral protein linked to the genome (VPg) of Turnip mosaic virus (TuMV; a member of the Potyvirus genus) in yeast 2-hybrid experiments. The interaction has also been demonstrated in vivo in brassica and a mutation in the interaction domain of the TuMV VPg completely abolished the interaction. In other examples of recessive plant resistance to potyviruses, it has been shown that certain natural mutations in the eIF(iso)4E or eIF4E genes results in the VPg no longer being able to bind to the proteins and it is assumed that this is the mechanism underlying the resistance, where lack of binding means the viruses cannot translate their RNA in to protein and hence cannot replicate. We have shown that broad-spectrum resistance to TuMV in Brassica rapa is controlled by two genes, one of which is recessive (retr01) and the other dominant (ConTR01). retr01 corresponds to eIF(iso)4E and ConTR01 corresponds to eIF4E. The student will utilise the yeast 2-hybrid system to determine whether the brassica eIF4E and eIF(iso)4E alleles conferring resistance to TuMV bind to the TuMV VPg. He / she will also investigate whether the protein from alleles of these genes at other loci, particularly the other loci of the Syngenta lines in to which the resistance is being introgressed, interact with the TuMV VPg. If as in other plant - potyvirus interactions, the presence or absence of interaction correlates with susceptibility and resistance respectively, the yeast 2-hybrid system and sequencing will be used to search for novel / superior alleles of eIF4E and eIF(iso)4E in a range of different brassica species with a view to identifying new opportunities to produce sources of resistance in these different brassica types. We have a large collection of over 200 TuMV isolates obtained from different plant species growing in many different parts of the world. Many of the isolates have been characterised in terms of their pathotype, serotype and genetic group. The VPg will be cloned from TuMV isolates from distinctly different genetic groups and their interaction with eIF4E and eIF(iso)4E alleles conferring resistance to TuMV will be determined in the yeast 2-hybrid system in order to investigate the potential durability of such resistance. Syngenta Netherlands was selected as the partner for the project as it is Syngenta's site responsible for vegetable breeding and has all the infrastructure and expertise to compliment the academic partner and take the project and materials forward for commercialisation, whilst providing the student with excellent industrial experience. The student will work with Syngenta Netherlands on the introgression of the broad-spectrum resistance to TuMV into commercial varieties of B. rapa using markers for the alleles of eIF(iso)4E and eIF4E that confer resistance to TuMV. They will verify the ability of the markers to discriminate the alleles conferring resistance from those alleles conferring susceptibility. He / she will also amplify and sequence the other copies of eIF4E and eIF(iso)4E present at other loci in the B. rapa lines in to which the resistance will be introgressed in order to determine whether they are likely to interfere with the resistance. The student will also phenotype plant lines produced during introgression of the resistance in to commercial B. rapa types, to determine whether they are resistant or susceptible to TuMV and thereby verify genotype determined using the markers. The susceptibility / resistance status of any brassica lines possessing potentially superior alleles of eIF4E and / or eIF(iso)4E will also be determined. With a four year studentship, there is sufficient time and flexibility for the student to develop their own ideas and approaches and pursue complimentary aspects to the project whilst achieving the goals outlined above.