Changes in mRNA Abundance within Heterodera schachtii-Infected Roots of Arabidopsis thaliana

Abstract

Gene expression changes in plant roots infected by plant-parasitic cyst nematodes are involved in the formation of nematode feeding sites. We analyzed mRNA abundance changes within roots of Arabidopsis thaliana during the early compatible interaction with Heterodera schachtii, the sugarbeet cyst nematode. Approximately 1,600 root sections, each containing a single parasitic nematode and its feeding site, and 1,600 adjacent, nematode-free root sections were excised from aseptic A. thaliana cultures 3 to 4 days after inoculation with H. schachtii. These tissue samples were termed infected and uninfected, respectively. Preparasitic nematodes were added to the uninfected tissue sample to maintain the nematode to plant tissue proportion. mRNA extracted from these two tissue samples was subjected to differential display analysis. Thirty-six cDNA clones corresponding to mRNA species with different abundance between both tissue samples were isolated. Of these clones, 24 were of A. thaliana origin and 12 were from H. schachtii. Differential display data predicted that the A. thaliana cDNA clones corresponded to 13 transcripts that were more abundant in the infected root sections and 11 transcripts that were more abundant in the uninfected root sections. H. schachtii cDNA clones were predicted to correspond to four transcripts that were more abundant in parasitic nematodes and to eight transcripts that were more abundant in preparasitic nematodes. In situ hybridization experiments confirmed the mRNA abundance changes in A. thaliana roots predicted by the differential display analyses for two A. thaliana clones.

Molecular Characterization of At5PTase1, an Inositol Phosphatase Capable of Terminating Inositol Trisphosphate Signaling,

“…A recent report detailing the identification of up-regulated mRNAs in nematode-infected Arabidopsis roots provides a physiological process in which At5PTase1 may be involved (Hermsmeier et al, 2000). As the At5PTase1 gene becomes expressed 68 times greater upon nematode infection, it is possible that signal termination via At5PTase1-catalyzed IP 3 breakdown is important for this plant-animal interaction.…”

Transcriptomic Analysis of Nematode Infestation

“…Frequently, whole root systems were used as a starting material for RNA isolation (Hermsmeier et al 1998 ;Mahalingam et al 1999 ;Vaghchhipawala et al 2001) . Furthermore, chances of identifying genes that were differentially regulated by nematode infections were improved by collecting tissues enriched for nematode feeding sites using approaches of preselecting infested tissues (for example, giant-cell or syncytial containing tissues) (Hermsmeier et al 2000 ;Vercauteren et al 2001) , by manually dissecting feeding sites or by microaspiration (Wieczorek et al 2006) . RNA isolated from such tissues was either immediately transcribed into cDNA or first enriched for feeding-site specific transcripts by subtractive hybridization.…”

“…However, unlike Lea genes, drought stress or ABA application did not induce the expression of the Lemmi9 gene in tomato. Hermsmeier et al (2000) examined early changes in transcript abundance in developing synctia induced by H. schachtii in Arabidopsis. Root segments with protruding juveniles were dissected 4 days after inoculation and compared to adjacent nematode-free tissues.…”

Nematode Infection Triggers the de Novo Formation of Unloading Phloem That Allows Macromolecular Trafficking of Green Fluorescent Protein into Syncytia

“…Gene expression profiling using differential display or DNA microarray techniques on H. schachtii-infected versus noninfected roots revealed 24 genes (Hermsmeier et al, 2000) or 128 genes (Puthoff et al, 2003) with altered transcript levels. These changes in gene expression could not, however, clearly be assigned to the syncytium but rather to the root zone containing this nematode-induced structure.…”