Project description:In order to study the similarities and differences in embryonic development between plant-parasitic nematodes and free-living nematodes, we performed RNA-seq on embryos of three plant-parasitic nematodes at a total of 11 stages from the single-cell stage to the J1 stage
Project description:Background: Epigenetic processes play an important role in the plant response to adverse environmental conditions. A role for DNA hypomethylation has recently been suggested in the pathogenic interaction between bacteria and plants, yet it remains unclear whether this phenomenon reflects a conserved and general plant immunity response. We therefore investigated the role of DNA methylation in the plant defence against damaging parasitic nematodes. Methods and results: Treatment of roots of rice (monocot plant) and tomato (dicot plant) by a nematode-associated molecular pattern (NAMP) from different parasitic nematodes revealed global DNA hypomethylation using ELISA based quantification, suggesting conservation among plants. Focusing on root-knot induced gall tissue in rice, the causal impact of hypomethylation on immunity was revealed by a significantly reduced plant susceptibility upon 5-Azacitidine treatment. Whole genome bisulfite sequencing revealed that hypomethylation was massively present in the CHH context, while absent for CpG or CHG nucleotide contexts. CHH hypomethylated regions were predominantly associated with gene promoter regions, which was not correlated with activated gene expression at the same time point, but rather showed a delayed effect on transcriptional gene activation. Finally, the relevance of CHH hypomethylation in plant defence was confirmed in rice mutants of the RNA-directed DNA methylation pathway (RdDM) and DDM1, which are known to be steering DNA methylation in CHH context. Conclusions: We demonstrated that DNA hypomethylation confers enhanced defence in rice towards root-parasitic nematodes and is likely to be part of the basal NAMP-triggered immunity response in plants.
Project description:Mobile small RNAs are an integral component of the arms race between plants and fungal parasites, and several studies suggest microRNAs could similarly operate between parasitic nematodes and their animal hosts. However, whether and how specific sequences are selected for export by parasites is unknown. Here we describe a specific Argonaute protein (exWAGO) that is secreted in extracellular vesicles (EVs) released by the gastrointestinal nematode Heligmosomodies bakeri, at multiple copies per EV. Phylogenetic and gene expression analyses demonstrate exWAGO is highly conserved and abundantly expressed in related parasites, including the human hookworm and proteomic analyses confirm this is the only Argonaute secreted by rodent parasites. In contrast, exWAGO orthologues in species from the free-living genus Caenorhabditis are highly diverged. By sequencing multiple small RNA libraries, we determined that the most abundant small RNAs released from the nematode parasite are not microRNAs but rather secondary small interfering RNAs (siRNAs) that are produced by RNA-dependent RNA Polymerases. We further identify distinct evolutionary properties of the siRNAs resident in free-living or parasitic nematodes versus those exported in EVs by the parasite and show that the latter are specifically associated with exWAGO. Together this work identifies an Argonaute protein as a mediator of RNA export and suggests rhabditomorph nematode parasites may have co-opted a novel nematode-unique pathway to communicate with their hosts.
Project description:Animal development is complex yet surprisingly robust. Animals may develop alternative phenotypes conditional on environmental changes. Under unfavorable conditions C. elegans larvae enter the dauer stage, a developmentally arrested, long-lived, and stress-resistant state. Dauer larvae of free-living nematodes and infective larvae of parasitic nematodes share many traits including a conserved endocrine signaling module (DA/DAF-12), which is essential for the formation of dauer and infective larvae. We speculated that conserved post-transcriptional regulatory mechanism might also be involved in executing the dauer and infective larvae fate. We used an unbiased sequencing strategy to characterize the microRNA (miRNA) gene complement in C. elegans, P. pacificus, and S. ratti. Our study raised the number of described miRNA genes to 257 for C. elegans, tripled the known gene set for P. pacificus to 362 miRNAs and is the first to describe miRNAs in a Strongyloides parasite. Moreover, we found a limited core set of 24 conserved miRNA families in all three species. Interestingly, our estimated expression fold changes between dauer vs. non-dauer stages and infective larvae vs. free-living stages reveal that despite the speed of miRNA gene set evolution in nematodes, homologous gene families with conserved ‘dauer-infective’ expression signatures are present. These findings suggest that common post-transcriptional regulatory mechanisms are at work and that the same miRNA families play important roles in developmental arrest as well as long-term survival in free-living and parasitic nematodes.
Project description:Animal development is complex yet surprisingly robust. Animals may develop alternative phenotypes conditional on environmental changes. Under unfavorable conditions C. elegans larvae enter the dauer stage, a developmentally arrested, long-lived, and stress-resistant state. Dauer larvae of free-living nematodes and infective larvae of parasitic nematodes share many traits including a conserved endocrine signaling module (DA/DAF-12), which is essential for the formation of dauer and infective larvae. We speculated that conserved post-transcriptional regulatory mechanism might also be involved in executing the dauer and infective larvae fate. We used an unbiased sequencing strategy to characterize the microRNA (miRNA) gene complement in C. elegans, P. pacificus, and S. ratti. Our study raised the number of described miRNA genes to 257 for C. elegans, tripled the known gene set for P. pacificus to 362 miRNAs and is the first to describe miRNAs in a Strongyloides parasite. Moreover, we found a limited core set of 24 conserved miRNA families in all three species. Interestingly, our estimated expression fold changes between dauer vs. non-dauer stages and infective larvae vs. free-living stages reveal that despite the speed of miRNA gene set evolution in nematodes, homologous gene families with conserved ‘dauer-infective’ expression signatures are present. These findings suggest that common post-transcriptional regulatory mechanisms are at work and that the same miRNA families play important roles in developmental arrest as well as long-term survival in free-living and parasitic nematodes. miRNA profiling in mixed and developmentally arrested stages (dauer/infective larvae) of nematodes by small RNA deep sequencing using Illumina GAII, Illumina HiSeq 2000, and ABI SOLiD.
Project description:The helminth Acanthocheilonema viteae serves as model organism for research on parasitic filarial nematodes. Total RNA secreted or excreted by 1500 adult female and male A. viteae over 3 weeks was isolated from culture media previously processed by differential ultracentrifugation, and subjected to miRNA sequencing.