DIFFERENTIAL IN-VITRO DNA-BINDING ACTIVITY TO A PROMOTER ELEMENT OF THE GN1 BETA-1,3-GLUCANASE GENE IN HYPERSENSITIVELY REACTING TOBACCO PLANTS

In a hypersensitive reaction to pathogen infection, expression of the beta-1,3-glucanase gn1 gene is induced in cells surrounding the necrot ic lesions. The 5'-flanking sequence of gn1 was examined to investigat e the molecular basis controlling activation of gene expression during this plant defense response. Studies on transgenic tobacco plants con taining gn1 promoter deletions fused to the beta-glucuronidase reporte r gene revealed the presence of negative and positive regulatory seque nces mediating both the level and the spatial distribution of gn1 expr ession. Promoter sequences to -138 bp were sufficient to confer increa sed gene expression around the necrotic lesions produced in response t o Pseudomonas syringae pv. syringae inoculation. It is demonstrated by electrophoretic mobility shift assays that nuclear proteins in both h ealthy and hypersensitively reacting tobacco leaves interact with DNA sequences within the regulatory elements identified. Among the binding sequences characterized, the promoter region extending from -250 to - 217 bp contained the DNA motif -GGCGGC- found to be conserved in most if not all promoters of genes encoding pathogenesis-related basic prot eins. The activity bound by this promoter sequence was stronger in hyp ersensitively responding tissues than in healthy untreated tobacco lea ves.