1b.Approach (from AD-416)
Using agrobacterium-mediated transient expression systems to investigate nematode CLE processing in planta.
Plant-parasitic cyst nematodes are known to secrete proteins that mimic the function of plant CLE signaling peptides to promote a successful infection. However, the mechanistic details of this process have yet to be elucidated. We have identified and functionally characterized CLE-like genes from several cyst nematode species including the two species of potato cyst nematodes. We have demonstrated that nematode CLEs can be trafficked by the function of the variable domain to the place outside plant cells where they can act as plant CLE mimics by interacting with membrane bound plant CLE receptors. We have further used a transient expression system to demonstrate the processing of nematode CLE in plants. The majority of the research in 2010 was focused on the identification of the processed and bioactive forms of nematode CLE peptides that function in plant cells and the initial investigation of plant receptors that perceive nematode CLE signals. By over-expression of a nematode CLE protein in potato roots coupled with proteomic analyses, we have determined that the bioactive forms of nematode CLEs share striking structural similarity to mature plant CLE peptides. Our results provide the direct evidence that nematode CLEs, once being delivered into host root cells, can be recognized by host cellular machinery to become mimics of plant CLE signals. Using genetic and biochemical approaches, we demonstrated that plant receptors of CLV2 and CRN are required for nematode CLE-mediated parasitism. We further identified that BAM1 and BAM2 are additional host receptors that can bind to processed nematode CLE peptides. These results suggested that multiple host receptors may be involved in perceiving nematode CLE peptides to facilitate nematode parasitism. Mimicry of host plant signals is an extraordinary adaptation by a plant parasite to have evolved the ability to reprogram host plant cells for its own benefit. A further study of this exciting example of molecular mimicry will discover novel strategies to bioengineer crops with robust resistance to pathogenic cyst nematodes.