Martin J. Wubben

Nematode effector proteins: an emerging paradigm of parasitism

Phytonematodes use a stylet and secreted effectors to modify host cells and ingest nutrients to support their growth and development. The molecular function of nematode effectors is currently the subject of intense investigation. In this review, we summarize our current understanding of nematode effectors, with a particular focus on proteinaceous stylet-secreted effectors of sedentary endoparasitic phytonematodes, for which a wealth of information has surfaced in the past 10 yr. We provide an update on the effector repertoires of several of the most economically important genera of phytonematodes and discuss current approaches to dissecting their function. Lastly, we highlight the latest breakthroughs in effector discovery that promise to shed new light on effector diversity and function across the phylum Nematoda.

Susceptibility to the Sugar Beet Cyst Nematode Is Modulated by Ethylene Signal Transduction in Arabidopsis thaliana

Previously, we identified Arabidopsis thaliana mutant rhd1-4 as hypersusceptible to the sugar beet cyst nematode Heterodera schachtii. We assessed rhd1-4 as well as two other rhd1 alleles and found that each exhibited, in addition to H. schachtii hypersusceptibility, decreased root length, increased root hair length and density, and deformation of the root epidermal cells compared with wild-type A. thaliana ecotype Columbia (Col-0). Treatment of rhd1-4 and Col-0 with the ethylene inhibitors 2-aminoethoxyvinylglycine and silver nitrate and the ethylene precursor 1-aminocyclopropane-1-carboxylic acid suggests that the rhd1-4 hypersusceptibility and root morphology phenotypes are the result of an increased ethylene response. Assessment of known ethylene mutants further support the finding that ethylene plays a role in mediating A. thaliana susceptibility to H. schachtii because mutants that overproduce ethylene (eto1-1, eto2, and eto3) are hypersusceptible to H. schachtii and mutants that are ethylene-insensitive (etr1-1, ein2-1, ein3-1, eir1-1, and axr2) are less susceptible to H. schachtii. Because the ethylene mutants tested show altered susceptibility and altered root hair density and length, a discrimination between the effects of altered ethylene signal transduction and root hair density on susceptibility was accomplished by analyzing the ttg and gl2 mutants, which produce ectopic root hairs that result in greatly increased root hair densities while maintaining normal ethylene signal transduction. The observed normal susceptibilities to H. schachtii of ttg and g12 indicate that increased root hair density, per se, does not cause hypersusceptibility. Furthermore, the results of nematode attraction assays suggest that the hypersusceptibility of rhd1-4 and the ethylene-overproducing mutant eto3 may be the result of increased attraction of H. schachtii-infective juveniles to root exudates of these plants. Our findings indicate that rhd1 is altered in its ethylene response and that ethylene signal transduction positively influences plant susceptibility to cyst nematodes.

Cyst Nematode Parasitism of Arabidopsis thaliana Is Inhibited by Salicylic Acid (SA) and Elicits Uncoupled SA-Independent Pathogenesis-Related Gene Expression in Roots

Compatible plant-nematode interactions involve the formation of an elaborate feeding site within the host root that requires the evasion of plant defense mechanisms by the parasite. Little is known regarding plant defense signaling pathways that limit nematode parasitism during a compatible interaction. Therefore, we utilized Arabidopsis thaliana mutants perturbed in salicylic acid (SA) biosynthesis or signal transduction to investigate the role of SA in inhibiting parasitism by the beet cyst nematode Heterodera schachtii. We determined that SA-deficient mutants (sid2-1, pad4-1, and NahG) exhibited increased susceptibility to H. schachtii. In contrast, SA-treated wild-type plants showed decreased H. schachtii susceptibility. The npr1-2 and npr1-3 mutants, which are impaired in SA signaling, also showed increased susceptibility to H. schachtii, whereas the npr1-suppressor mutation sni1 showed decreased susceptibility. Constitutive pathogenesis-related (PR) gene-expressing mutants (cpr1 and cpr6) did not show altered susceptibility to H. schachtii; however, constitutive PR gene expression was restricted to cpr1 shoots with wild-type levels of PR-1 transcript present in cpr1 roots. Furthermore, we determined that H. schachtii infection elicits SA-independent PR-2 and PR-5 induction in wild-type roots, while PR-1 transcript and total SA levels remained unaltered. This was in contrast to shoots of infected plants where PR-1 transcript abundance and total SA levels were elevated. We conclude that SA acts via NPR1 to inhibit nematode parasitism which, in turn, is negatively regulated by SNI1. Our results show an inverse correlation between root basal PR-1 expression and plant susceptibility to H. schachtii and suggest that successful cyst nematode parasitism may involve a local suppression of SA signaling in roots.

Transcript analysis of parasitic females of the sedentary semi-endoparasitic nematode Rotylenchulus reniformis.

Rotylenchulus reniformis, the reniform nematode, is a sedentary semi-endoparasitic nematode capable of infecting >300 plant species, including a large number of crops such as cotton, soybean, and pineapple. In contrast to other economically important plant-parasitic nematodes, molecular genetic data regarding the R. reniformis transcriptome is virtually nonexistant. Herein, we present a survey of R. reniformis ESTs that were sequenced from a sedentary parasitic female cDNA library. Cluster analysis of 2004 high quality ESTs produced 123 contigs and 508 singletons for a total of 631 R. reniformis unigenes. BLASTX analyses revealed that 39% of all unigenes showed similarity to known proteins (E<or=1.0e-04). R. reniformis genes homologous to known parasitism genes were identified and included beta-1,4-endoglucanase, fatty acid- and retinol-binding proteins, and an esophageal gland cell-specific gene from Heterodera glycines. Furthermore, a putative ortholog of an enzyme involved in thiamin biosynthesis, thought to exist solely in prokaryotes, fungi, and plants, was identified. Lastly, 114 R. reniformis unigenes orthologous to RNAi-lethal Caenorhabditis elegans genes were discovered. The work described here offers a glimpse into the transcriptome of a sedentary semi-endoparasitic nematode which (i) provides the transcript sequence data necessary for investigating engineered resistance against R. reniformis and (ii) hints at the existance of a thiamin biosynthesis pathway in an animal.

Molecular characterization and temporal expression analyses indicate that the MIC (Meloidogyne Induced Cotton) gene family represents a novel group of root-specific defense-related genes in upland cotton (Gossypium hirsutum L.)

The molecular events underlying the resistance of Upland cotton (Gossypium hirsutum L.) to the root-knot nematode (RKN) are largely unknown. In this report, we further characterize the previously identified MIC3 gene including the identification of 14 related MIC cDNAs in nematode-infected roots of allotetraploid cotton that show >85% identity with MIC3. A time-course analysis of RKN infection in resistant and susceptible cotton lines showed that maximum MIC transcript accumulation occurred immediately prior to the phenotypic manifestation of resistance. MIC expression was not induced by mechanical wounding or by virulent reniform nematode infection. MIC expression was undetectable in cotton leaves undergoing a hypersensitive response to Xanthomonas campestris. A time-course analysis of defense gene expression (PR10, ERF5, CDNS, LOX1, POD4, POD8) in resistant and susceptible cotton roots showed that RKN infection specifically elicits the induction of MIC in resistant roots and not other common defense-signaling pathways. These results suggest that cotton resistance to RKN involves novel defense-signaling pathways and further supports the idea that the MIC genes are intimately involved in this resistance response and represent a group of root-specific defense-related genes in cotton.

The genome of the cotton bacterial blight pathogen Xanthomonas citri pv. malvacearum strain MSCT1

Xanthomonas citri pv. malvacearum is a major pathogen of cotton, Gossypium hirsutum L.. In this study we report the complete genome of the X. citri pv. malvacearum strain MSCT1 assembled from long read DNA sequencing technology. The MSCT1 genome is the first X. citri pv. malvacearum genome with complete coding regions for X. citri pv. malvacearum transcriptional activator-like effectors. In addition functional and structural annotations are presented in this study that will provide a foundation for future pathogenesis studies with MSCT1.