The fire blight resistance of apple clones in which DspE-interacting proteins are silenced

The disease specific gene, dspE (syn. dspA) of Erwinia amylovora, encodes a 198 kD pathogenicity effector that is essential for development of the fire blight disease in plants. Yeast two-hybrid and in vitro protein pull-down assays demonstrated that DspE interacts physically and specifically with four similar leucine-rich-repeat, receptor-like serine/theonine kinases from apple. Genes encoding the four DspE-interacting proteins of Malus (DIPM genes) are conserved in all hosts of E. amylovora tested, but not in tested non-host plants. The interaction between the DIPM's and DspE is thought to be involved in disease development. Sense sequences of ca. 400 bp from non-conserved regions of each DIPM gene, with homology among each other of <50%, were used to make constructs to be transformed into fire blight susceptible apple cultivars with the aim of silencing the DIPM genes and thereby preventing interaction with DspE. In addition, three constructs containing the four ca. 400 bp sequences of the individual genes in tandem, a full length sense sequence of one gene, and a "hairpin" sequence of that gene were made. All seven silencing sequences were transferred into 'Galaxy' apple cultivar by Agrobacterium inoculation of leaf pieces. Transgenic lines with all constructs were recovered. Assays of the transgenic plants for net expression of the target DIPM's by RT-PCR for their mRNA's have shown evidence of partial to complete silencing of some of the DIPM's in some transgenic lines. Evaluation for resistance to fire blight by inoculation of shoots with E. amylovora strain Ea273 indicated that some lines with silencing had increased resistance. Resistance due to silencing of a native apple gene(s) is likely to be more acceptable to regulators, growers and consumers than the use of heterologous.