Project description:Purpose: The aim of this study is to investigate the role of H3K9 methylation in the development of syncytia induced by the beet cyst nematode (BCN, Heterodera schachtii). For this, ChIP-seq analysis was conducted in root tissues of Col-0 wild-type versus single suvh5 and suvh6 mutant plants and under BCN-infected and noninfected conditions. Methods: Methods: Col-0 wild-type and suvh5 and suvh6 mutant plants were grown and 10-day-old plants were inoculated with 100 second-stage juvenile (J2) H. schachtii at 10 days old. Five days post inoculation, root tissues were collected from infected and noninfected plants in triplicate. ChIP DNA was precipitated using the H3k9me2 antibody (Abcam) and prepared into libraries. Then, high throughput ChIP DNA sequencing was performed using the Illumina NovoSeq 6000. Raw reads were trimmed of adapter sequences and overrepresented reads. Then, high quality, paired-end reads were aligned to the Arabidopsis thaliana reference genome (TAIR10.28) using Bowtie2. Finally, histone peaks were called using MACS2 for the identification of genes associated with differentially dimethylated H3K9 in wild-type versus mutant roots under BCN-infected and noninfected conditions. Results and Conclusions: Under noninfected conditions, 511 and 143 H3K9me2 peaks were enriched in Col-0 compared to suvh5 and suvh6, respectively. The majority of these H3K9me2 peaks are located within TEs, which is consistent with previous findings that H3K9me2 corresponds to the repression of detrimental TEs. Our results suggest, however, that H3K9me2 adopts a new role in response to BCN. BCN-infected Col-0 had 791 H3K9me2 peaks compared to noninfected Col-0, 743 of which are located in the promoter or body of protein-coding genes. These peaks are associated with 1180 protein-coding genes. Similarly, suvh5 and suvh6 single mutants had 353 and 3106 H3K9me2 peaks compared to BCN-infected Col-0, respectively. The majority of these peaks are located in protein-coding genes and are associated with 558 and 4594 protein-coding genes, respectively. Interestingly, about 65% of peaks enriched in the BCN-infected wild type overlapped with peaks depleted in the suvh6 mutant, indicating that SUVH6 contributes to the majority of H3K9me2 during BCN parasitism.

Project description:Purpose: The aim of this study is to investigate the role of H3K9 methylation in the development of syncytia induced by the beet cyst nematode (BCN, Heterodera schachtii). For this, differential gene expression analysis was performed in root tissues of Col-0 wild-type and suvh5 and suvh6 single mutants under BCN-infected and noninfected conditions. Methods: Col-0 wild-type and suvh5 and suvh6 mutant plants were grown and 10-day-old plants were inoculated with 100 second-stage juvenile (J2) H. schachtii at 10 days old. Five days post inoculation, root tissues were collected from infected and noninfected plants in triplicate. Total RNA was extracted, mRNA libraries were prepared, and then high throughput RNA sequencing was performed using the Illumina NovoSeq 6000. High quality, paired-end reads were then aligned to the Arabidopsis thaliana reference genome (TAIR10.28) using STAR and uniquely mapped reads were counted using HTSeq. Finally, differentially expressed genes (DEGs) between the mutant lines and wild-type plants under non-infected and infected conditions were identified using DESeq package. Results and conclusions: At least 20 million reads were uniquely mapped to the Arabidopsis reference genome with an average length of about 300 bp for paired-end reads. A total of 1057 and 3390 DEGs were identified for suvh5 and suvh6 versus Col-0, respectively, under noninfected conditions. Upon infection of BCN, Col-0 had 4566 DEGs compared to the noninfected wild type. BCN-infected single mutants suvh5 and suvh6 had 2487 and 846 DEGs compared to BCN-infected Col-0, respectively. Of these DEGs, 1277 and 592 or about 51% and 70% are also differentially expressed in BCN-infected Col compared to noninfected Col-0, respectively, and 324 of the 1277 and 592 DEGs are common to BCN-infected suvh5 and suvh6 compared to the control. DEGs common to BCN-infected Col-0 and/or suvh5, and suvh6 are involved in stress and defense responses, primary and secondary metabolism, signal transduction, cell wall organization and other cellular processes. This suggests that SUVH5 and SUVH6 play a major role in transcriptome reprogramming to facilitate cellular, molecular, and metabolic processes involved in the compatible plant-nematode interaction.

Project description:BackgroundSugar beet (Beta vulgaris subsp. vulgaris) is an economically important crop that provides nearly one third of the global sugar production. The beet cyst nematode (BCN), Heterodera schachtii, causes major yield losses in sugar beet and other crops worldwide. The most effective and economic approach to control this nematode is growing tolerant or resistant cultivars. To identify candidate genes involved in susceptibility and resistance, the transcriptome of sugar beet and BCN in compatible and incompatible interactions at two time points was studied using mRNA-seq.ResultsIn the susceptible cultivar, most defense-related genes were induced at 4 dai while suppressed at 10 dai but in the resistant cultivar Nemakill, induction of genes involved in the plant defense response was observed at both time points. In the compatible interaction, alterations in phytohormone-related genes were detected. The effect of exogenous application of Methyl Jasmonate and ET-generator ethephon on susceptible plants was therefore investigated and the results revealed significant reduction in plant susceptibility. Genes putatively involved in the resistance of Nemakill were identified, such as genes involved in phenylpropanoid pathway and genes encoding CYSTM domain-containing proteins, F-box proteins, chitinase, galactono-1,4-lactone dehydrogenase and CASP-like protein. Also, the transcriptome of the BCN was analyzed in infected root samples and several novel potential nematode effector genes were found.ConclusionsOur data provides detailed insights into the plant and nematode transcriptional changes occurring during compatible and incompatible interactions between sugar beet and BCN. Many important genes playing potential roles in susceptibility or resistance of sugar beet against BCN, as well as some BCN effectors with a potential role as avr proteins were identified. In addition, our findings indicate the effective role of jasmonate and ethylene in enhancing sugar beet defense response against BCN. This research provides new molecular insights into the plant-nematode interactions that can be used to design novel management strategies against BCN.

Project description:Cyst nematodes are devastating plant parasites that become sedentary within plant roots and induce the transformation of normal plant cells into elaborate feeding cells with the help of secreted effectors, the parasitism proteins. These proteins are the translation products of parasitism genes and are secreted molecular tools that allow cyst nematodes to infect plants.We present here the expression patterns of all previously described parasitism genes of the soybean cyst nematode, Heterodera glycines, in all major life stages except the adult male. These insights were gained by analyzing our gene expression dataset from experiments using the Affymetrix Soybean Genome Array GeneChip, which contains probeset sequences for 6,860 genes derived from preparasitic and parasitic H. glycines life stages. Targeting the identification of additional H. glycines parasitism-associated genes, we isolated 633 genes encoding secretory proteins using algorithms to predict secretory signal peptides. Furthermore, because some of the known H. glycines parasitism proteins have strongest similarity to proteins of plants and microbes, we searched for predicted protein sequences that showed their highest similarities to plant or microbial proteins and identified 156 H. glycines genes, some of which also contained a signal peptide. Analyses of the expression profiles of these genes allowed the formulation of hypotheses about potential roles in parasitism. This is the first study combining sequence analyses of a substantial EST dataset with microarray expression data of all major life stages (except adult males) for the identification and characterization of putative parasitism-associated proteins in any parasitic nematode.We have established an expression atlas for all known H. glycines parasitism genes. Furthermore, in an effort to identify additional H. glycines genes with putative functions in parasitism, we have reduced the currently known 6,860 H. glycines genes to a pool of 788 most promising candidate genes (including known parasitism genes) and documented their expression profiles. Using our approach to pre-select genes likely involved in parasitism now allows detailed functional analyses in a manner not feasible for larger numbers of genes. The generation of the candidate pool described here is an important enabling advance because it will significantly facilitate the unraveling of fascinating plant-animal interactions and deliver knowledge that can be transferred to other pathogen-host systems. Ultimately, the exploration of true parasitism genes verified from the gene pool delineated here will identify weaknesses in the nematode life cycle that can be exploited by novel anti-nematode efforts.

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Interplay between gene expression, histone modifications, and DNA methylation mediated by miR778 during beet cyst nematode parasitism of Arabidopsis

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