Poplar CLE peptides stimulating ectomycorrhizal symbiosis uncovered by genome-wide identification of ectomycorrhizal-responsive small secreted peptides

Abstract

Plants small secreted peptides (SSPs) regulate root development, immunity and symbiotic relationships in herbaceous plants. These processes are equally important for establishing functional ectomycorrhizal associations in trees. While fungal SSPs involved in ectomycorrhizal establishment have been identified, the role of plant SSPs remains largely unexplored. Although thousands of SSPs were predicted in plant genomes, their small size and high sequence divergence hinder their accurate automated annotations. To address this issue, we combined de novo gene prediction with family-specific motif search to identify 1,053 SSPs from 21 symbiosis-related families in the genomes of two ectomycorrhizal tree species: Populus trichocarpa and Quercus robur. Nearly half of these SSPs are transcriptionally regulated during ectomycorrhizal symbiosis with various fungal partners. Functional assays of selected Populus CLE peptides, a SSP family known to repress arbuscular mycorrhizal and rhizobial symbioses, revealed that five enhanced ectomycorrhizal roots formation. Unlike CLEs involved in the autoregulation of arbuscular mycorrhizal and rhizobial symbioses, these peptides belonged to clades associated with meristem activity. Their activity did not increase lateral root number but inhibited adventitious root growth, suggesting their role in promoting ectomycorrhizal root organogenesis. These findings demonstrate that CLE peptides promote, rather than repress, ectomycorrhizal symbiosis. This functional divergence from their roles in other symbioses suggests that poplar co-opted a distinct set of SSPs to support ectomycorrhizal development. Our results expand the understanding of host tree contributions to ectomycorrhizal development and identify a set of candidate SSPs for future functional studies, thereby highlighting a new layer of regulation in tree-fungal mutualism.