The TIR-NB-LRR pair DSC1 and WRKY19 contributes to basal immunity of Arabidopsis to the root-knot nematode Meloidogyne incognita

BackgroundRoot-knot nematodes transform vascular host cells into permanent feeding structures to withdraw nutrients from the host plant. Ecotypes of Arabidopsis thaliana can display large quantitative variation in susceptibility to the root-knot nematode Meloidogyne incognita, which is thought to be independent of dominant major resistance genes. However, in an earlier genome-wide association study of the interaction between Arabidopsis and M. incognita we identified a quantitative trait locus harboring homologs of dominant resistance genes but with minor effect on susceptibility to the M. incognita population tested.ResultsHere, we report on the characterization of two of these genes encoding the TIR-NB-LRR immune receptor DSC1 (DOMINANT SUPPRESSOR OF Camta 3 NUMBER 1) and the TIR-NB-LRR-WRKY-MAPx protein WRKY19 in nematode-infected Arabidopsis roots. Nematode infection studies and whole transcriptome analyses using the Arabidopsis mutants showed that DSC1 and WRKY19 co-regulate susceptibility of Arabidopsis to M. incognita.ConclusionGiven the head-to-head orientation of DSC1 and WRKY19 in the Arabidopsis genome our data suggests that both genes may function as a TIR-NB-LRR immune receptor pair. Unlike other TIR-NB-LRR pairs involved in dominant disease resistance in plants, DSC1 and WRKY19 most likely regulate basal levels of immunity to root-knot nematodes

Skewed X-inactivation is common in the general female population

X-inactivation is a well-established dosage compensation mechanism ensuring that X-chromosomal genes are expressed at comparable levels in males and females. Skewed X-inactivation is often explained by negative selection of one of the alleles. We demonstrate that imbalanced expression of the paternal and maternal X-chromosomes is common in the general population and that the random nature of the X-inactivation mechanism can be sufficient to explain the imbalance. To this end, we analyzed blood-derived RNA and whole-genome sequencing data from 79 female children and their parents from the Genome of the Netherlands project. We calculated the median ratio of the paternal over total counts at all X-chromosomal heterozygous single-nucleotide variants with coverage ≥10. We identified two individuals where the same X-chromosome was inactivated in all cells. Imbalanced expression of the two X-chromosomes (ratios ≤0.35 or ≥0.65) was observed in nearly 50% of the population. The empirically observed skewing is explained by a theoretical model where X-inactivation takes place in an embryonic stage in which eight cells give rise to the hematopoietic compartment. Genes escaping X-inactivation are expressed from both alleles and therefore demonstrate less skewing than inactivated genes. Using this characteristic, we identified three novel escapee genes (SSR4, REPS2, and SEPT6), but did not find support for many previously reported escapee genes in blood. Our collective data suggest that skewed X-inactivation is common in the general population. This may contribute to manifestation of symptoms in carriers of recessive X-linked disorders. We recommend that X-inactivation results should not be used lightly in the interpretation of X-linked variants

Skewed X-inactivation is common in the general female population

X-inactivation is a well-established dosage compensation mechanism ensuring that X-chromosomal genes are expressed at comparable levels in males and females. Skewed X-inactivation is often explained by negative selection of one of the alleles. We demonstrate that imbalanced expression of the paternal and maternal X-chromosomes is common in the general population and that the random nature of the X-inactivation mechanism can be sufficient to explain the imbalance. To this end, we analyzed blood-derived RNA and whole-genome sequencing data from 79 female children and their parents from the Genome of the Netherlands project. We calculated the median ratio of the paternal over total counts at all X-chromosomal heterozygous single-nucleotide variants with coverage ≥10. We identified two individuals where the same X-chromosome was inactivated in all cells. Imbalanced expression of the two X-chromosomes (ratios ≤0.35 or ≥0.65) was observed in nearly 50% of the population. The empirically observed skewing is explained by a theoretical model where X-inactivation takes place in an embryonic stage in which eight cells give rise to the hematopoietic compartment. Genes escaping X-inactivation are expressed from both alleles and therefore demonstrate less skewing than inactivated genes. Using this characteristic, we identified three novel escapee genes (SSR4, REPS2, and SEPT6), but did not find support for many previously reported escapee genes in blood. Our collective data suggest that skewed X-inactivation is common in the general population. This may contribute to manifestation of symptoms in carriers of recessive X-linked disorders. We recommend that X-inactivation results should not be used lightly in the interpretation of X-linked variants

The root-knot nematode effector MiMSP32 targets host 12-oxophytodienoate reductase 2 (OPR2) to regulate plant susceptibility

To establish persistent infections in host plants, herbivorous invaders, such as root-knot nematodes, must rely on effectors for suppressing damage-induced jasmonate-dependent host defenses. However, at present, the effector mechanisms targeting the biosynthesis of biologically active jasmonates to avoid adverse host responses are unknown.Using yeast-two-hybrid, in planta co-immunoprecipitation, and mutant analyses, we identified 12-oxophytodienoate reductase 2 (OPR2) as an important host target of the stylet-secreted effector MiMSP32 of the root-knot nematode Meloidogyne incognita. MiMSP32 has no informative sequence similarities with other functionally annotated genes but was selected for the discovery of novel effector mechanisms based on evidence of positive, diversifying selection.OPR2 catalyzes the conversion of a derivative of 12-oxophytodienoate to jasmonic acid (JA) and operates parallel to 12-oxophytodienoate reductase 3 (OPR3), which controls the main pathway in the biosynthesis of jasmonates. We show that MiMSP32 targets OPR2 to promote parasitism of M. incognita in host plants independent of OPR3-mediated JA biosynthesis.Artificially manipulating the conversion of the 12-oxophytodienoate by OPRs increases susceptibility to multiple unrelated plant invaders. Our study is the first to shed light on a novel effector mechanism targeting this process to regulate susceptibility of host plant

Circulating TP53 mutations are associated with early tumor progression and poor survival in pancreatic cancer patients treated with FOLFIRINOX

Background: Biomarkers predicting treatment response may be used to stratify pancreatic ductal adenocarcinoma (PDAC) patients for therapy. The aim of this study was to identify circulating tumor DNA (ctDNA) mutations that associate with tumor progression during FOLFIRINOX chemotherapy, and overall survival (OS). Methods: Circulating cell-free DNA was analyzed with a 57 gene next-generation sequencing panel using plasma samples of 48 PDAC patients of all disease stages. Patients received FOLFIRINOX as initial treatment. Chemotherapy response was determined on CT scans as disease control (n = 30) or progressive disease (n = 18) within eight cycles of FOLFIRINOX, based on RECIST 1.1 criteria. Results: Detection of a TP53 ctDNA mutation before start of FOLFIRINOX [odds ratio (OR) 10.51, 95% confidence interval (CI) 1.40–79.14] and the presence of a homozygous TP53 Pro72Arg germline variant (OR 6.98, 95% CI 1.31–37.30) were predictors of early tumor progression during FOLFIRINOX in multivariable analysis. Five patients presented with the combination of a TP53 ctDNA mutation before start of FOLFIRINOX and the homozygous Pro72Arg variant. All five patients showed progression during FOLFIRINOX. The combination of the TP53 mutation and TP53 germline variant was associated with shorter survival (median OS 4.4 months, 95% CI 2.6–6.2 months) compared with patients without any TP53 alterations (median OS 13.0 months, 95% CI 8.6–17.4 months). Conclusion: The combination of a TP53 ctDNA mutation before start of FOLFIRINOX and a homozygous TP53 Pro72Arg variant is a promising biomarker, associated with early tumor progression during FOLFIRINOX and poor OS. The results of this exploratory study need to be validated in an independent cohort

The expression of Gr-VAP1 coincides with host invasion and migration of <i>Globodera rostochiensis</i>.

<p>The expression of <i>Gr-VAP1</i>, as shown by semi-quantitative reverse transcription PCR, is highly up-regulated in the migratory stages of <i>G. rostochiensis</i> (ppJ2, J2, and males (♂)), while it declines after initiation of the permanent feeding site in the sedentary juvenile stages (J3 and J4, and adult females (♀). Changes in expression of <i>Gr-VAP1</i> were assessed using the constitutively expressed <i>cAMP-</i>dependent protein kinase (<i>cAMP</i>) gene in <i>G. rostochiensis</i> as reference. Reactions using uninfected tomato roots as template (Root) and without reverse transcriptase (-RT) were included as controls.</p

Ectopic venom allergen-like proteins from cyst and root-knot nematodes selectively suppress defense-related programmed cell death.

<p>(A) Agroinfiltration assays in <i>Nicotiana benthamiana</i> showing the transient co-expression in the apoplast of cell death inducing elicitin <i>INF1</i> of <i>Phytophthora infestans</i> and venom allergen-like proteins from <i>G. rostochiensis</i> (Gr-VAP1), <i>H. schachtii</i> (Hs-VAP1 and Hs-VAP2), and <i>Meloidogyne incognita</i> (Mi-VAP1). Co-expressions with the corresponding empty binary vector (EV) and green fluorescent protein (GFP) were included as controls. (B and C) Transient co-expression of receptor-like proteins Cf-4 and Cf-9 from tomato and their cognate elicitors Avr4 and Avr9 from <i>C. fulvum</i> with venom allergen-like proteins and controls as described above. Photographs were taken 4 days post infiltration for INF1, and 7 days post infiltration for Cf-4/Avr4 and Cf9/Avr9. The bars represent the mean number of events in which cell death suppression was observed for a total of 60 inoculation spots over 5 biological replicates (with standard error of mean). Different letters indicate a significant difference when using <i>P</i>-value <0.05 (in an ANOVA).</p

Ectopic venom allergen-like proteins suppress basal immunity in <i>Arabidopsis thaliana</i>.

<p>(A) Heterologous expression of the venom allergen-like protein Gr-VAP1 from <i>G. rostochiensis</i>, and Hs-VAP1 and Hs-VAP2 from <i>Heterodera schachtii</i> in the apoplast of transgenic Arabidopsis lines enhances their susceptibility to <i>H. schachtii</i>. Two independent transgenic lines per construct (-A and -B) were compared with corresponding transgenic line harboring the T-DNA of the empty vector (EV) and wild type <i>A. thaliana</i> (Col-0). Bars represent mean number of nematodes per plants with standard errors of the means. Letters indicate statistical significance when using <i>P</i>-value <0.05 as threshold. (B) Ectopic Hs-VAP1 and Hs-VAP2 enhance development of disease symptoms of fungal and oomycete pathogens in leaves of transgenic Arabidopsis lines. Pictures show symptoms on leaves inoculated with <i>Botrytis cinerea</i>, <i>Plectosphaerella cucumeria</i>, and two isolates of <i>Phytophthora brassicae</i>, or mock inoculated. (C) Ectopic Hs-VAP1 and Hs-VAP2 suppress seedling growth response of Arabidopsis to the immunogenic peptide flg22. Bars represent mean root length of transgenic lines with standard error of mean after 10 days in the presence or absence of 10 µM flg22.</p

Ten most down-regulated transcripts by ectopic venom allergen-like proteins in Arabidopsis.

<p>Differentially expressed genes in transgenic <i>Arabidopsis thaliana</i> overexpressing <i>Hs-VAP1</i> and <i>Hs-VAP2</i> relative to the corresponding transgenic empty vector control plants (for full lists see <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004569#ppat.1004569.s008" target="_blank">S1 Table</a> and <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1004569#ppat.1004569.s009" target="_blank">S2 Table</a>).</p><p>* The fold change (FC) is calculated as standardized log2-transformed counts per million relative to transgenic plants harboring the corresponding empty vector.</p><p>Ten most down-regulated transcripts by ectopic venom allergen-like proteins in Arabidopsis.</p