1.1.30
DIVERGENT EVOLUTION OF THREE CLUSTERS OF PLANT RESISTANCE GENE HOMOLOGUES

M PARNISKE1, BBH WULFF1, G BONNEMA2 and JDG JONES1

1The Sainsbury Laboratory, John lnnes Centre, Colney Lane, Norwich NR4 7UH, UK; 2University of Wageningen, Wageningen, The Netherlands

Background and objectives
Hcr9 genes (homologues of Cladosporium fulvum resistance gene Cf-9) encode plant resistance against tomato leaf mould by the recognition of different avirulence determinants in the pathogen. The genes reside in complex loci carrying multiple genes and encode predicted membrane-bound proteins with extracytoplasmic leucine-rich repeats (LRRs). In order to unravel the mechanisms involved in generating novel recognition specificities, we genetically mapped, isolated and sequenced multiple members of the gene family.

Materials and methods
Three loci of clustered Hcr9 genes were identified on the short arm of chromosome 1 of tomato. YAC and phage clones were isolated for the two loci mapping distal (Northern Lights) and proximal (Southern Cross) to the Cf-4/9 locus (Milky Way). The nucleotide sequence of two Hcr9s from Southern Cross and six Hcr9s from Northern Lights, including intergenic regions, was determined.

Results and conclusions
Sequence comparisons between Hcr9s from all three loci confirmed the earlier finding that novel sequence variants are generated by sequence exchange between gene family members. The entire cluster organization of Southern Cross and Milky Way is very similar, and the sequences are closely related to each other, suggesting that these clusters are the result of a recent duplication event. Alternatively, the similarity might be caused by ongoing sequence exchange between these loci by ectopic recombination. In contrast, Northern Lights is more distantly related, indicating that this locus resulted from a more ancient duplication event and/or is exchanging sequences with the other loci at a lower rate. The organization of Hcr9 genes in three clusters, of which at least one is evolving independently, might serve as a mechanism to increase sequence divergence in the gene family.

References
1. Parniske MKE, Hammond-Kosack C, Goistein CM et al., 1997. Cell 91, 821-832.
2. Thomas CM, Jones DA, Parniske M et al., 1997. Plant Cell 9, 2209-2224.