Soybean transporter AAT (Rhg1) abundance increases along the nematode migration path and impacts vesiculation and ROS

Rhg1 (Resistance to Heterodera glycines 1) mediates soybean (Glycine max) resistance to soybean cyst nematode (SCN; H. glycines). Rhg1 is a 4-gene, ∼30-kb block that exhibits copy number variation, and the common PI 88788-type rhg1-b haplotype carries 9 to 10 tandem Rhg1 repeats. Glyma.18G022400 (Rhg1-GmAAT), 1 of 3 resistance-conferring genes at the complex Rhg1 locus, encodes the putative amino acid transporter AAT(Rhg1) whose mode of action is largely unknown. We discovered that AAT(Rhg1) protein abundance increases 7- to 15-fold throughout root cells along the migration path of SCN. These root cells develop an increased abundance of vesicles and large vesicle-like bodies (VLB) as well as multivesicular and paramural bodies. AAT(Rhg1) protein is often present in these structures. AAT(Rhg1) abundance remained low in syncytia (plant cells reprogrammed by SCN for feeding), unlike the Rhg1 α-SNAP protein, whose abundance has previously been shown to increase in syncytia. In Nicotiana benthamiana, if soybean AAT(Rhg1) was present, oxidative stress promoted the formation of large VLB, many of which contained AAT(Rhg1). AAT(Rhg1) interacted with the soybean NADPH oxidase GmRBOHG, the ortholog of Arabidopsis thaliana RBOHD previously found to exhibit upregulated expression upon SCN infection. AAT(Rhg1) stimulated reactive oxygen species (ROS) generation when AAT(Rhg1) and GmRBOHG were co-expressed. These findings suggest that AAT(Rhg1) contributes to SCN resistance along the migration path as SCN invades the plant and does so, at least in part, by increasing ROS production. In light of previous findings about α-SNAP(Rhg1), this study also shows that different Rhg1 resistance proteins function via at least 2 spatially and temporally separate modes of action.