Identification of HcrVf2 as an apple scab resistance gene and characterisation of HcrVf control sequences

by Eve Silfverberg-Dilworth

Abstract

Apple scab caused by the fungal pathogen Venturia inaequalis is one of the most damaging diseases affecting apple production, especially in areas with high spring and summer rainfall. Natural resistance to apple scab is found in many wild Malus species. Vf (Venturia floribunda) apple scab resistance derived from the wild apple Malus floribunda 821 is the most studied and widely used source of apple scab resistance. Inheritance studies show that Vf is due to a single locus, as is Vf virulence in V. inaequalis demonstrating a gene-for-gene interaction. Gene-for-gene interactions are proposed to involve a plant receptor protein (encoded by a plant resistance gene) that recognises a pathogen elicitor (encoded by a pathogen avirulence gene). A cluster of resistance gene homologues was recently identified in the region of the Vf locus (named HcrVf genes). They are predicted to encode transmembrane proteins with an extracellular receptor domain.
This thesis describes the functional analysis of these genes via gene inactivation in a scab-resistant apple cultivar and gene transfer to a scab-susceptible cultivar. Gene silencing, using an antisense HcrVf sequence, in the apple scab resistant cultivar Florina appears to lead to plant death, especially following plant wounding. The R gene homologue HcrVf2 was successfully transferred to the scab-susceptible cultivar Gala via Agrobacterium-mediated transformation. Inoculations with spore suspensions of V. inaequalis showed both in vitro and in vivo that this gene is involved in scab resistance. Inoculations with both Vf virulent and Vf avirulent races of V. inaequalis demonstrated that this gene is involved in recognition of individual pathogen races.
It has been suggested that developmental regulation of some of these R gene homologues (HcrVf genes) may occur. The identification of their upstream, (5’) promoter sequences, responsible for gene expression is described in this thesis. These promoters can now be used for the further investigation of the control of HcrVf gene expression.
Identification of avirulence genes or elicitors from V. inaequalis and further investigation of the other resistance gene homologues of the Vf region is now required. This will help to further clarify whether genes other than HcrVf2, the gene analysed in this thesis, are involved in race-specific interactions between Vf and V. inaequalis.

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