SAGE Analisis of Syncytia

Several hundred species of nematodes live as obligate biotrophic plant parasites that attack mainly the roots of a variety of plants causing worldwide crop losses of $ 100 billion per year. Sedentary endoparasitic nematodes enter the plant root and establish specific feeding structures which allow them to obtain all their nutrients from the host plant. The feeding structure of root-knot nematodes consists of several giant cells whereas the cyst nematodes induce the formation of a syncytium which is initiated from a single root cell and expands by incorporating up to several hundred cells by local cell wall dissolutions. We are studying the compatible interaction between Arabidopsis thaliana and the beet cyct nematode Heterodera schachtii. Arabidopsis plants can be grown and infected with nematodes in vitro and their thin and translucent roots make it possible to directly observe the nematodes by light microscopy. This will allow us to use microaspiration to obtain pure syncytium material which will then be used to extract total RNA and to analyse the transcriptome of syncytia using SAGE (Serial Analysis of Gene Expression) technology. We will compare the transcriptome of young, expanding syncytia and mature syncytia. This will allow us to identify genes that are needed for growth of the syncytium, for instance those coding for cell wall degrading enzymes, and genes that are important for supporting the parasite with nutrients. The expression of interesting genes identified by this SAGE analysis will be further studied by RT-PCR and in situ RT-PCR. Their expression will also be compared with that in giant cells induced by Meloidogyne incognita.

Several hundred species of nematodes live as obligate biotrophic plant parasites that attack mainly the roots of a variety of plants causing worldwide crop losses of $ 100 billion per year. Sedentary endoparasitic nematodes enter the plant root and establish specific feeding structures which allow them to obtain all their nutrients from the host plant. The feeding structure of root-knot nematodes consists of several giant cells whereas the cyst nematodes induce the formation of a syncytium which is initiated from a single root cell and expands by incorporating up to several hundred cells by local cell wall dissolutions. We are studying the compatible interaction between Arabidopsis thaliana and the beet cyct nematode Heterodera schachtii. Arabidopsis plants can be grown and infected with nematodes in vitro and their thin and translucent roots make it possible to directly observe the nematodes by light microscopy. This will allow us to use microaspiration to obtain pure syncytium material which will then be used to extract total RNA and to analyse the transcriptome of syncytia using SAGE (Serial Analysis of Gene Expression) technology. We will compare the transcriptome of young, expanding syncytia and mature syncytia. This will allow us to identify genes that are needed for growth of the syncytium, for instance those coding for cell wall degrading enzymes, and genes that are important for supporting the parasite with nutrients. The expression of interesting genes identified by this SAGE analysis will be further studied by RT-PCR and in situ RT-PCR. Their expression will also be compared with that in giant cells induced by Meloidogyne incognita.