Identification of a stylet-secreted effector protein family as a core component of root-knot nematode feeding tubes

Proteins secreted from a mouth stylet of sedentary plant-parasitic root-knot nematodes self-polymerize to form a unique feeding tube structure within host cells modified into giant feeding cells by the nematode. Feeding tubes have essential functions as they complex with the host endomembrane system for nutrient uptake to sustain parasitism. Despite their significance, they remain one of the least understood aspects of nematode parasitism of plants. Their small size and location within giant-cells deeply embedded within galls encasing adult females has prohibited studies to isolate and discern their molecular composition. Here, we developed a protocol for the isolation and semipurification of root-knot nematode feeding tubes from giant-cell cytoplasm of several host plant species to provide a unique view of these structures at the light and scanning electron microscopy level revealing previously undescribed features of their structure. Our methods allowed for the isolation and solubilization of sufficient quantities of enriched feeding tubes enabling a comparative proteome analysis across host species that identified proteins with an increased likelihood to function in feeding tube formation. A comparison across root-knot nematode species further narrowed candidates to a conserved class of secretory proteins that specifically localized within secretory granules of the dorsal gland of adult females and in feeding tubes formed within host cell cytoplasm to unequivocally demonstrate these proteins as core components of feeding tubes. Our finding gives scientists a look into the protein composition of feeding tubes opening the door to a better understanding of their structure and function in nematode parasitism.