GENE-FOR-GENE INTERACTIONS: THE ROLE OF AVIRULENCE GENES IN PATHOGENICITY AND RACE-SPECIFIC RESISTANCE
PJGM DE WIT, MHAJ JOOSTEN, R LAUGE, R ROTH, R LUDERER, N WESTERINK, G HONEE, M KOOMAN-GERSMANN, F LAURENT, RAL VAN DER HOORN, CF DE JONG, P VOSSEN and R WEIDE
Department of Phytopathology, Wageningen Agricultural University, Binnenhaven 9, 6709 PD Wageningen, The Netherlands
Background and objectives
Various plant pathogens, including viruses, bacteria and fungi, interact with their hosts in a gene-for-gene manner. The gene-for-gene model implies recognition of products of avirulence (avr) genes by products of matching resistance (R) genes, which would act as ligands and receptors, respectively. Recognition of specific elicitors often leads to a hypersensitive response (HR) and activation of other defence responses. Many R genes and avr genes have been cloned in recent years. There is much homology between the various R genes, but hardly any homology between various avr genes. The intrinsic functions of avr genes are often not known. However, it has been demonstrated that some avr genes play a role in pathogenicity. These encode proteins with known functions such as replicase for viruses. For most bacteria and fungi, however, a role in pathogenicity for avr genes could only be demonstrated by decrease in pathogenicity or virulence after avr gene replacement in the pathogen. They include the bacterial avr genes avrA, avrB, avrE, avrBs2 and avrRPM1, and the fungal avr genes nipl, Ecp1 and Ecp2. Several groups have been searching for receptors of specific elicitors in order to prove or disprove that they represent products of R genes. A short overview of these studies will be presented.
Results and conclusions
In our own laboratory we have been searching for receptors of the Cladosporium fulvum elicitors AVR9 and AVR4. For AVR9, high-affinity binding sites with similar Kds have been found in plasma membranes of solanaceous and other plant species. No binding sites were detected in Arabidopsis. So far, we did not find increased binding of 125I-AVR9 to plasma membranes of Cf-g transgenic tobacco and no binding to Cf-9 transgenic Arabidopsis, indicating that AVR9 does not bind to the CF9 protein, or does bind with such a low affinity that we could not detect it with the methods presently available. From an alanine scan of AVR9, mutant peptides were found with lower, similar or higher HR-inducing activity compared to wild-type AVR9, while some mutants showed hardly any HR-inducing activity on leaves of Cf9 tomato genotypes. A positive correlation between HR-inducing activity and binding affinity was found for these mutants, suggesting a role for the high-affinity binding site in HR-induction in Cf9 plants. Cell suspensions generated from Cfg tomato genotypes do not respond to AVR9, while cell suspensions from Cf9 transgenic tobacco do respond with a specific pH shift and an oxidative burst after treatment with the various AVR9 peptides. Replacement of the cysteine residues in AVR4 by alanine demonstrated that all eight cysteine residues form disuifide bridges, of which two could be assigned. The AVR4 elicitor protein can be produced in Pichia pastoris, which enables studies on the global structure of AVR4 by NMR and binding assays to plasma membranes of Cf4 tomato genotypes and other plant species. We have performed a successful search for plants responding with HR among accessions of tomato and other solanaceous species, after exposure to ECP1 or ECP2. The matching genes, designated Cf-ECP1 and Cf-ECP2, respectively, are expected to be durable as they are targeted against pathogenicity factors of C. fulvum.