Abstract
Protein disulfide isomerase (PDI) is a multifunctional enzyme that catalyzes rate-limiting reactions such as disulfide bond formation, isomerization, and reduction. There is some evidence that indicates that PDI is also involved in host-pathogen interactions in plants. In this study, we show that the rice root-knot nematode, Meloidogyne graminicola, has evolved a secreted effector, MgPDI2, which is expressed in the subventral esophageal glands and up-regulated during the early parasitic stage of M. graminicola. Purified recombinant MgPDI2 functions as an insulin disulfide reductase and protects plasmid DNA from nicking. As an effector, MgPDI2 contributes to nematode parasitism. Silencing of MgPDI2 by RNA interference in the pre-parasitic second-stage juveniles (J2s) reduced M. graminicola multiplication and also increased M. graminicola mortality under H2O2 stress. In addition, an Agrobacterium-mediated transient expression assay found that MgPDI2 caused noticeable cell death in Nicotiana benthamiana. An intact C-terminal region containing the first catalytic domain (a) with an active motif (Cys-Gly-His-Cys, CGHC) and the two non-active domains (b and b′) is required for cell death induction in N. benthamiana. This research may provide a promising target for the development of new strategies to combat M. graminicola infections.
Highlights
Protein disulfide isomerase (PDI) is a member of the thioredoxin superfamily and is a multifunctional enzyme that catalyzes rate-limiting reactions such as disulfide bond formation, isomerization, and reduction in the endoplasmic reticulum (ER) [1,2]
We found that MgPDI2 encodes a typical PDI in the endoparasitic nematode M. graminicola
These results indicated that MgPDI2 might play a role in M. graminicola virulence
Summary
Protein disulfide isomerase (PDI) is a member of the thioredoxin superfamily and is a multifunctional enzyme that catalyzes rate-limiting reactions such as disulfide bond formation, isomerization, and reduction in the endoplasmic reticulum (ER) [1,2]. A PDI gene (PpPDI1) from Phytophthora parasitica contributes to plant infection and causes necrosis of tobacco leaves when transiently expressed in Nicotiana benthamiana [11]. In order to determine if MgPDI2 can work as a pathogenicity factor that induces cell death in N. benthamiana, an Agrobacterium-mediated transient expression assay in N. benthamiana leaves was performed. Transient expression of MgPDI2 caused noticeable cell death in N. benthamiana 48 h after Agro-infiltration, while the empty vector pGD-eGFP construct did not induce cell death (Figure 7). Western blotting indicated that the MgPDI2:pGD-eGFP fusion protein was expressed in the leaves of N. benthamiana, thereby supporting a role for MgPDI2 in inducing host plant necrosis. The results of our Western blot confirmed that these mutated fusion proteins were expressed in the leaves of N. benthamiana (Figure 8B), while the presence of numerous bands might be due to the poor antibody specificity.
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