5.4.3S
INFECTION-INDUCED EXPRESSION OF THE AVIRULENCE GENE Avr9 IN TRANSGENIC CF9 TOMATO PLANTS CONFERS RESISTANCE TO FUNGAL PATHOGEN ATTACK

MH STUIVER2, G HONTE1, R WEIDE1, H TIGFLAAR2, LS MELCHERS2 and PJGM DE WIT1

1Department of Phytopathology, Agricultural University, Wageningen, Binnenhaven 9, 6709 PD, Wageningen, The Netherlands; 2Zeneca-MOGEN, PO Box 628, 2300 AP Leiden, The Netherlands

Background and objectives
The most effective defence response of plants against pathogen attack is probably the hypersensitive response (HRI) which comprises a localized collapse of tissue surrounding the infection site. The genetic basis of the HR-mediated disease resistance in tomato to the fungal pathogen Cladosporium fulvum is described by the gene-for-gene model [1]. Tomato plants carrying the Cf-9 resistance gene are resistant against attack of C. fulvum races which contain the avirulence gene Avr9. The Avr9 gene encodes a pre-pro-protein of 63 amino acids which after secretion is converted by proteases into a mature elicitor peptide of 28 amino acids. The Cf-9 gene encodes a putative membrane-anchored extracytoplasmic glycoprotein which consists of 28 impect leucine-rich repeats. Injection of the AVR9 peptide into leaves of tomato plants carrying the Cf-9 resistance gene results in HR confined to the infected area. Constitutive expression of the Avr9 gene in transgenic Cf-9 plants results in delayed growth, necrosis and complete plant death. We have tried to employ components of this system to generate a broad-spectrum pathogen resistance.

Materials and methods
Promoter activity of the defence-related gene gst1 from potato is induced upon pathogen attack. Constructs were designed containing the Avr9 coding region under the transcriptional control of a gst1 promoter fragment [2]. These constructs were used for Agrobacterium-mediated transformation of Cf-9 leaf discs which resulted in transgenic plants showing normal phenotype.

Results and conclusions
Transgenic plants were selfed and progeny plants were used for inoculation with races of C. fulvum lacking the Avr9 gene. Several lines showed infection-induced HR and resistance to C. fulvum. Microscopical observations showed restricted fungal growth in transgenic leaves, while in wild-type Cf-9 leaves the growth was abundant. Fungal resistance was stably inherited and co-segregated with the presence of the Avr9 transgene. Lines that exhibit increased resistance towards C. fulvum also have increased levels of resistance towards other fungi, including Oidium lycopersici, the causal agent of powdery mildew in tomato. Also, in this infection HR events and restriction of fungal growth were clearly observed in the transgenic lines.

References
1. De Wit PJGM, 1992. Annual Review of Phytopathology 30, 391-418.
2. Strittmatter et al., 1996. Molecular Plant-Microbe Interactions 9, 68-73.