Nathalie Barthels

Characterization of a pathogen-induced potato catalase and its systemic expression upon nematode and bacterial infection

We have isolated a cDNA encoding a catalase (Cat2St) by differential screening of a cDNA library constructed from potato roots infected with the cyst nematode Globodera pallida. Expression analysis confirmed the local induction of Cat2St and showed that it was highest at the adult stage of the parasite. It also revealed that Cat2St was induced in uninfected roots, stems, and leaves of infected plants. Localized and systemic induction of Cat2St was also observed upon root-knot nematode (Meloidogyne incognita) and root bacteria (Erwinia carotovora, Corynebacterium sepedonicum) infections. Based on sequence and expression analysis, Cat2St was found to belong to the recently described class II of dicotyledonous catalases, suggesting that these catalase isoforms could also be pathogen induced. Plant-parasitic nematodes are known to induce, in the roots of their hosts, highly metabolic feeding cells that function as nutritional sinks. Whereas the local induction of Cat2St is probably a consequence of an oxidative stress of metabolic nature, the systemic induction of Cat2St shows striking similarities with the induction of systemic acquired resistance (SAR) genes. The possible role of catalase in compatible plant-pathogen interactions is discussed.

Isolation of the LEMMI9 gene and promoter analysis during a compatible plant-nematode interaction

Plant-endoparasitic root-knot nematodes feed on specialized giant cells that they induce in the vascular cylinder of susceptible plants. Although it has been established that a number of plant genes change their expression pattern during giant cell differentiation, virtually no data are available about the mechanisms involved in that change. One possibility is differential promoter recognition by the transcription factor(s) responsible for the expression of specific genes. We have isolated and characterized a genomic clone from tomato containing the promoter region of LEMMI9, one of the few plant genes that have been reported to be highly expressed in galls (predominantly in giant cells). The analysis of transgenic potato plants carrying a LEMMI9 promoter-beta glucuronidase (GUS) fusion has demonstrated that the tomato promoter was activated in Meloidogyne incognita-induced galls in a heterologous system. We have located putative regulatory sequences in the promoter and have found that nuclear proteins from the galls formed specific DNA-protein complexes with the proximal region of the LEMMI9 promoter. The nuclear protein-binding sequence mapped to a region of 111 bp immediately upstream from the TATA box. This region contains a 12-bp repeat possibly involved in the formation of DNA-protein complexes, which might be related to the LEMMI9 transcriptional activation in the giant cells.

Arabidopsis Thaliana as a Model Host Plant to Study Molecular Interactions with Root-Knot and Cyst Nematodes

Arabidopsis thaliana is a small cruciferous plant that is considered as being a “botanical Drosophila” (Whyte, 1946) for a number of reasons recently reviewed by Meyerowitz (1989). Thanks to its small size and ability to grow well at room temperature both in vitro and in the greenhouse, it is easy and cheap to maintain large populations of Arabidopsis plants under laboratory conditions. Arabidopsis has a short generation time: approximately 3 months from seed to seed. It is extremely prolific and its seeds are very small (about 50,000 seeds fit in a 1.5-ml Eppendorf tube). These features make A. thaliana highly suited for classical genetic approaches.

Integration of nematode-responsive regulatory sequences from Arabidopsis thaliana into nematode control strategies

While for many decades fundamental research on sedentary nematode-plant parasitism encompassed mainly the ultrastructural and physiological aspects of the interaction, more recently also the area of molecular biology has started to be explored. Although the puzzle of the underlying gene regulation is far from complete, the available knowledge already enables us to evaluate the effectiveness of several envisaged genetic engineering strategies to obtain crops with nematode-resistant properties. Both antifeeding structure and antinematode approaches are considered and involve a gene complex in which a specific nematode-inducible promoter is responsible for the local production of a selected gene product aimed at inhibiting proper feeding cell and/or nematode development. Effective nematode control will greatly depend on the choice of both components in such a complex. To identify nematode-inducible plant-regulatory sequences, we have used the model system Arabidopsis thaliana. We report here on several transgenic lines tagged in promising regulatory sequences and discuss their potential in the scope of engineering nematode resistance into plants.

Plant gene expression in nematode-induced feeding sites and strategies for engineering nematode control.

Efforts by several research teams to gather information on how feeding cells of sedentary endoparasitic nematodes are molecularly supported evolve more and more toward a comprehension of the involved plant gene expression and regulation. Effective genetically engineered nematode control can be achieved by using a combination of a specific nematode-inducible promoter and the destructive effect of a selected gene product targeted toward the nematode or the feeding cell. Feeding site-expressed genes are currently being catalogued. The expression of many plant genes with known function and pattern is being monitored by using reporter gene systems (e.g. beta-glucuronidase) to study the potential role of these genes in the interaction between the nematode and its host. Specific searches including the screening of cDNA libraries and, more recently, the differential display technique allowed the identification of differentially expressed sequences. Promoter traps have been successfully used to isolate nematode-responsive regulatory regions and deletion studies are aimed to obtain more refined nematode-responsive elements. So far, linkage of a tagged promoter with a native gene could not often be proven, but such a correspondence might not be necessary for engineering nematode control. Because the plants gene regulation was not developed as such to serve nematodes, strict confinement of gene activity to feeding structures is not always feasible and therefore the so-called two-component system might be useful for its counterbalancing effect on eventual undesired leakiness. The more the catalogue broadens, the more possibilities it will create to find the most suitable and promising promoter-gene combination to fight nematode attack.