Laccaria bicolor MiSSP8 is a small-secreted protein decisive for the establishment of the ectomycorrhizal symbiosis

The ectomycorrhizal symbiosis is a predominant tree-microbe interaction in forest ecosystems sustaining tree growth and health. Its establishment and functioning implies a long-term and intimate relationship between the soil-borne fungi and the roots of trees. Mycorrhiza-induced Small Secreted Proteins (MiSSPs) are hypothesized as keystone symbiotic genes, required to set up the symbiosis by modifying the host metabolism and/or building the symbiotic interfaces. Since L. bicolor MiSSP8 is the third most highly induced MiSSPs in symbiotic tissues and it is expressed in fruiting bodies, we report its functional study by analysing phenotypes of L. bicolor MiSSP8-RNAi mutants and MiSSP8 in planta subcellular localization. The MiSSP8-RNAi knockdown mutants are impaired in their mycorrhization ability with Populus, with the lack of fungal mantle and Hartig net development. When expressed heterologously in planta, MiSSP8 shows a punctate localization that partially colocalize with plasmodesmata. Moreover, MiSSP8 displays a fungal-specific repetitive motif, present in proteins from both saprotrophic and ectomycorrhizal fungi. MiSSP8 is a Laccaria-specific small-secreted protein required for early stages of ectomycorrhizal symbiosis. Our data suggest that MiSSP8 has a possible dual role: one in basic fungal biology regulating hyphal aggregation and another in planta, related to its plasmodesmata co-localization.