The root‐knot nematode effector MiPDI1 targets a stress‐associated protein (SAP) to establish disease in Solanaceae and Arabidopsis

Abstract

Large amounts of effectors are secreted by the oesophageal glands of plant-parasitic nematodes, but their molecular mode of action remains largely unknown. We characterized a Meloidogyne incognita protein disulphide isomerase (PDI)-like effector protein (MiPDI1) that facilitates nematode parasitism. In situ hybridization showed that MiPDI1 was expressed specifically in the subventral glands of M. incognita. It was significantly upregulated during parasitic stages. Immunolocalization demonstrated MiPDI1 secretion in planta during nematode migration and within the feeding cells. Host-induced silencing of the MiPDI1 gene affected the ability of the nematode to infect the host, whereas MiPDI1 expression in Arabidopsis increased susceptibility to M. incognita, providing evidence for a key role of MiPDI1 in M. incognita parasitism. Yeast two-hybrid, bimolecular fluorescence complementation and coimmunoprecipitation assays showed that MiPDI1 interacted with a tomato stress-associated protein (SlSAP12) orthologous to the redox-regulated AtSAP12, which plays an important role in plant responses to abiotic and biotic stresses. SAP12 silencing or knocking out in Nicotiana benthamiana and Arabidopsis increased susceptibility to M. incognita. Our results suggest that MiPDI1 acts as a pathogenicity factor promoting disease by finetuning SAP-mediated responses at the interface of redox signalling, defence and stress acclimation in Solanaceae and Arabidopsis.

The Meloidogyne incognita Nuclear Effector MiEFF1 Interacts With Arabidopsis Cytosolic Glyceraldehyde-3-Phosphate Dehydrogenases to Promote Parasitism

“…Microarray analysis on Arabidopsis gapc mutants showed that several genes encoding for enzymes regulating reactive oxygen species (ROS) homeostasis, such as peroxidases, catalases, or superoxide dismutases, had altered expression (Rius et al, 2008), and here, using a RT-qPCR approach, we demonstrated a differential expression of defense-and stress-associated genes in abp39 and abx27 mutants, both mutant lines showing strong constitutive expression of AtPR1a and AtPDF1.2a genes. While several RKN effectors were shown to be responsible for host defense suppression (Quentin et al, 2013;Vieira and Gleason, 2019), very few have had their host target identified, i.e., MiMIF2 (Zhao et al, 2019), MiPDI1 (Zhao et al, 2020), Mg16820 (Naalden et al, 2018), MgMO237 (Chen et al, 2018), and MjTTL5 (Lin et al, 2016). This work adds MiEFF1 to the list of RKN effectors modulating host immunity and confirms that defense-and stress-related proteins are key targets of effectors, particularly those of nematodes, during disease establishment.…”

“…The M. incognita MiEFF1 (Minc17998; GenBank MW345915) and MiEFF18 (Minc18636; GenBank KX907770) coding sequences (CDS) lacking the signal peptide, the A. thaliana AtUSP (At3g53990), AtGAPC1 (At3g04120), AtGAPC2 (At1g13440), and AtSmD1b (At4g02840), the AtUSP and AtGAPC2 promoters, the SV40 T antigen and the murin P53 sequences were amplified by PCR with specific primers (Supplementary Table 1) and inserted into the pDON207 donor vector. The entry clones were recombined with pK7WGF2 (p35S:eGFP-GW), pK7WGR2 (p35S:RFP-GW), pKGWFS7 (GWpromoter:eGFP-GUS), BiFC vectors (pAM-35SS:GW-YFPc, pAM-35SS:GW-YFPn, pAM-35SS:YFPc-GW, and pAM-35SS:YFPn-GW) or Y2H vectors, pGBK-GW, pB27-GW, or pP6-GW (Karimi et al, 2002;Caillaud et al, 2009;Zhao et al, 2020), with Gateway technology (Invitrogen). All constructs were sequenced (GATC Biotech) and used to transform Agrobacterium tumefaciens strain GV3101 or Saccharomyces cerevisiae strains AH109, L40ΔGal4, or Y187.…”

“…The whole sequences encoding the AtUSP, AtGAPC1, and AtGAPC2 were cloned into the pP6 prey vectors, which were used to transform yeast strain Y187. The pairwise mating experiments with Y187 (matα) and L40ΔGal4 (mata) yeast strains were performed as previously described (Zhao et al, 2020).…”

“…To investigate whether AtGAPC1 and AtGAPC2 manipulation by MiEFF1 could affect Arabidopsis immunity, we measured, using RT-qPCR, the expression of genes involved in antioxidative functions (AtCDS2) and abiotic (AtADH1 and AtSAP12) or biotic (AtPR1a, AtPDF1.2a, and AtPR4) stress responses (Zhao et al, 2020) in abp39 and abx27 mutants. In both mutants, AtPR1a and AtPDF1.2a, regulators of salicylic acid (SA)-and ethylene-and jasmonate (JA)-mediated defense responses, respectively, showed a strong constitutive expression, while AtPR4, encoding an ethylene-responsive PR protein, was repressed in both lines (Figure 5).…”

“…Meloidogyne incognita secretes hundreds of effectors into host plants (Bellafiore et al, 2008;Wang et al, 2012). A role in parasitism has been demonstrated for very few of these effectors, and the target in the plant has been identified for only 10 of these proteins (Mejias et al, 2019(Mejias et al, , 2020Zhao et al, 2020). RKN effectors often target multifunctional proteins regulating key biological processes and conserved in all eukaryotes.…”