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Subcellular localization of resistance-associated AAPermeaseRhg1 in response to soybean cyst nematode infection

Shaojie HAN: University of Wisconsin-Madison, Department of Plant Pathology

<div>The <em>Rhg1</em> locus mediates economically valuable resistance of soybean to soybean cyst nematode. One of the three genes that contributes to the resistance encoded at the complex <em>Rhg1</em> locus encodes a putative amino acid permease (AAPermease<sub>Rhg1</sub>). How this AAPermease<sub>Rhg1</sub> mediates plant resistance to cyst nematodes is largely unknown. Here, we elucidated the subcellular location of AAPermease<sub>Rhg1</sub> during SCN infection. Using a custom-raised antibody to track the native protein, we performed transmission electron microscopy immunogold subcellular localization assays in resistant and susceptible roots samples. Samples were examined prior to infection and at time points 3 and 7 days after SCN infection. We also evaluated the cellular location by subcellular fractionation and by immunofluorescence using a wide-field confocal microscope. Upon root invasion, SCN juveniles (J2) move intracellularly, disrupting the cells through which they pass. We observed an increased abundance of small vesicles in penetrated root cells. AAPermease <sub>Rhg1</sub> signal was often associated with these small vesicles. The overall abundance of AAPermease<sub>Rhg1</sub> was up 7- to 15-fold compared to adjacent non-penetrated cells, while in general AAPermease<sub>Rhg1</sub> remained a low-abundance transmembrane protein in non-infected roots and in syncytia. These findings suggest that AAPermease<sub>Rhg1</sub> may contribute to SCN resistance via a unique localization change, directly surrounding the pathogen as it infects the plant.</div>