A mechanistic understanding of community ecology requires addressing the challenge of nonadditive effects of multispecies interactions, which requires the integration of ecological and molecular complexity— i.e., moving beyond pairwise ecological interaction studies and ‘gene-at-a-time’ approach to mechanism. Using the tripartite mutualism between arbuscular mycorrhizal fungi, nitrogen-fixing bacteria, and a model legume (Medicago truncatula), we investigate the consequences of multispecies mutualisms for the structure and function of genome-wide differential coexpression networks for the first time. Coexpression analysis identifies groups of genes (“coexpression modules”) whose expression change in concert, giving us a multivariate picture of molecular changes hosts and symbionts experience in response to multispecies interactions. We related changes in these complex molecular phenotypes to changes in plant performance in a multi-mutualism context and identify how the presence of third party mutualists impact joint expression across the symbiotic boundary (i.e., how rhizobia impacts coexpression of genes that reside in host plants with those that reside in the mycorrhizal fungi). We also placed the set of previously-identified candidate genes whose expression is affected additively or nonadditively by multiple mutualists in a coexpression network context to determine if these genes play central or peripheral roles in networks of gene interactions.
Results/Conclusions
We find extensive evidence for multispecies mutualisms affecting the coexpression networks across the genomes of participating species. First, multispecies mutualisms substantially changed coexpression network structure of 18 host plant gene modules and 22 mycorrhizal fungi gene modules, indicating that third-party mutualists (rhizobia) can cause significant rewiring of plant and fungal molecular networks. Second, we found the structure of the majority of coexpression modules that explained variation in plant performance were interactively affected by rhizobia and fungi, suggesting multiple mutualist changes in coexpression that may underlie performance changes. Third, across the symbiosis boundary we identified sets of plant and mycorrhizal genes whose coexpression structure was unique to the multiple mutualist context, and potentially coupled host-symbiont responses to rhizobia. Fourth, candidate genes previously-identified as nonadditively affected by multiple mutualists were highly connected in the coexpression networks and had 94% greater network centrality than additive genes, suggesting that nonadditive genes may be key players in the widespread transcriptomic responses to multispecies mutualisms. Our results show multispecies interactions have substantial consequences for molecular interactions in host plants, microbes, and across symbiotic boundaries and emphasizes the value for future work taking an integrative molecular and ecological approach to community interactions.