Summary The infection of potato (Solanum tuberosum) leaves with the late blight pathogen Phytophthora infestans, or treatment with fungal elicitor, leads to the massive accumulation of pathogenesis-related (PR) proteins in the extracellular leaf space. The most abundant of these proteins was purified to apparent homogeneity and identified as a new, basic member of the PR-1 family of defence proteins, designated PR-1b. Antibodies raised against the protein and a cDNA isolated by differential screening were used to study the temporal and spatial patterns of PR-1b protein and mRNA distribution in healthy and infected potato tissues. PR-1b was present in old leaves and at low levels also in the carpels of flowers. In leaves, strong accumulation of PR-1b mRNA and protein occurred in response to infection by the oomycete pathogen Phytophthora infestans or the bacterial pathogen Pseudomonas syringae pv. maculicola. PR-1b mRNA and protein accumulation was clearly initiated at the infection site, but a delayed and sustained accumulation was also observed in neighbouring, uninfected leaves of potato plants. Tissue- and cell type-specific expression of PR-1b was analysed by immunohistochemical and in situ RNA hybridization techniques. Appreciable amounts of PR-1b protein and mRNA were localized in epidermal cells, guard cells of the stomata, glandular trichomes, crystal idioblasts, and cells of the vascular system of infected leaves. However, no significant differences in the amounts and distribution patterns of PR-1b could be observed between compatible and incompatible interactions of potato and Phytophthora infestans, indicating that PR-1b expression is not involved in determining cultivar/race-specific resistance in potato.
Chitinases are ubiquitous proteins that occur in all plants in multiple isoforms. We have isolated the ChtC2 gene encoding an unusual, basic (class I) chitinase from potato ( Solanum tuberosum L.). In contrast to other chitinase genes, ChtC2 is not activated by infection, but rather constitutively expressed in leaves and stems where it is restricted to epidermal cells. Sequence analysis revealed a number of potential regulatory elements in the promoter, but most striking was the presence of a 319-bp direct repeat located between -333 and -968 upstream of the transcription start site. For a functional analysis, a 1,322-bp promoter fragment and two 5' deletions of 782 bp and 162 bp in length were translationally fused to the beta-glucuronidase (GUS) reporter gene and used for transient expression studies by particle bombardment. All promoter constructs conferred expression of GUS activity in different epidermal cell types of potato leaves. Expression in parenchyma cells of the leaf mesophyll was not detectable with any of the ChtC2 gene promoter constructs, in contrast to the pattern observed with the 35S promoter from cauliflower mosaic virus. The epidermis-specific expression of the reporter gene was confirmed using transgenic potato plants containing the fusion of the entire ChtC2 promoter with the GUS reporter. Histochemical analysis indicated that the promoter was only active in epidermal cells of leaves.
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