The ectomycorrhizal fungus Pisolithus microcarpus encodes a microRNA involved in cross-kingdom gene silencing during symbiosis

Small RNAs (sRNAs) are known to regulate pathogenic plant–microbe interactions. Emerging evidence from the study of these model systems suggests that microRNAs (miRNAs) can be translocated between microbes and plants to facilitate symbiosis. The roles of sRNAs in mutualistic mycorrhizal fungal interactions, however, are largely unknown. In this study, we characterized miRNAs encoded by the ectomycorrhizal fungus Pisolithus microcarpus and investigated their expression during mutualistic interaction with Eucalyptus grandis. Using sRNA sequencing data and in situ miRNA detection, a novel fungal miRNA, Pmic_miR-8, was found to be transported into E. grandis roots after interaction with P. microcarpus. Further characterization experiments demonstrate that inhibition of Pmic_miR-8 negatively impacts the maintenance of mycorrhizal roots in E. grandis, while supplementation of Pmic_miR-8 led to deeper integration of the fungus into plant tissues. Target prediction and experimental testing suggest that Pmic_miR-8 may target the host NB-ARC domain containing transcripts, suggesting a potential role for this miRNA in subverting host signaling to stabilize the symbiotic interaction. Altogether, we provide evidence of previously undescribed cross-kingdom sRNA transfer from ectomycorrhizal fungi to plant roots, shedding light onto the involvement of miRNAs during the developmental process of mutualistic symbioses.

A Transcriptomic Atlas of the Ectomycorrhizal Fungus Laccaria bicolor

Trees are able to colonize, establish and survive in a wide range of soils through associations with ectomycorrhizal (EcM) fungi. Proper functioning of EcM fungi implies the differentiation of structures within the fungal colony. A symbiotic structure is dedicated to nutrient exchange and the extramatricular mycelium explores soil for nutrients. Eventually, basidiocarps develop to assure last stages of sexual reproduction. The aim of this study is to understand how an EcM fungus uses its gene set to support functional differentiation and development of specialized morphological structures. We examined the transcriptomes of Laccaria bicolor under a series of experimental setups, including the growth with Populus tremula x alba at different developmental stages, basidiocarps and free-living mycelium, under various conditions of N, P and C supply. In particular, N supply induced global transcriptional changes, whereas responses to P supply seemed to be independent from it. Symbiosis development with poplar is characterized by transcriptional waves. Basidiocarp development shares transcriptional signatures with other basidiomycetes. Overlaps in transcriptional responses of L. bicolor hyphae to a host plant and N/C supply next to co-regulation of genes in basidiocarps and mature mycorrhiza were detected. Few genes are induced in a single condition only, but functional and morphological differentiation rather involves fine tuning of larger gene sets. Overall, this transcriptomic atlas builds a reference to study the function and stability of EcM symbiosis in distinct conditions using L. bicolor as a model and indicates both similarities and differences with other ectomycorrhizal fungi, allowing researchers to distinguish conserved processes such as basidiocarp development from nutrient homeostasis.

Ectomycorrhizal fungus supports endogenous rhythmic growth and corresponding resource allocation in oak during various below- and aboveground biotic interactions

Endogenous rhythmic growth (ERG) is displayed by many tropical and some major temperate tree species and characterized by alternating root and shoot flushes (RF and SF). These flushes occur parallel to changes in biomass partitioning and in allocation of recently assimilated carbon and nitrogen. To address how biotic interactions interplay with ERG, we cross-compared the RF/SF shifts in oak microcuttings in the presence of pathogens, consumers and a mycorrhiza helper bacterium, without and with an ectomycorrhizal fungus (EMF), and present a synthesis of the observations. The typical increase in carbon allocation to sink leaves during SF did not occur in the presence of root or leaf pathogens, and the increase in nitrogen allocation to lateral roots during RF did not occur with the pathogens. The RF/SF shifts in resource allocation were mostly restored upon additional interaction with the EMF. Its presence led to increased resource allocation to principal roots during RF, also when the oaks were inoculated additionally with other interactors. The interactors affected the alternating, rhythmic growth and resource allocation shifts between shoots and roots. The restoring role of the EMF on RF/SF changes in parallel to the corresponding enhanced carbon and nitrogen allocation to sink tissues suggests that the EMF is supporting plants in maintaining the ERG.

Austroboletus brunneisquamus (Boletaceae, Boletales), a New Ectomycorrhizal Fungus from a Tropical Rainforest, China

Austroboletusbrunneisquamus (Boletaceae/Boletales), an ectomycorrhizal fungus, is described as a new species from a tropical rainforest in China based on morphological and molecular evidence. It is morphologically characterized by a subtomentose pileal surface when young, which cracks into areolae, having large, pale brown and brown to dark brown scales, a stipe with yellowish brown reticulation, basidiospores measuring (11–)12–14.5(–15) × 6–8(–8.5) μm, with fine cristate to subreticulate ornamentation, and a pileipellis in the form of a cutis. A detailed description, color photographs of fresh basidiomata, and line drawings of microscopic features of the new species are presented.

The ectomycorrhizal basidiomycete Laccaria bicolor releases a GH28 polygalacturonase that plays a key role in symbiosis establishment

In ectomycorrhiza, root penetration and colonization of the intercellular space by symbiotic hyphae is thought to rely on the mechanical force that results from hyphal tip growth, enhanced by the activity of secreted cell-wall-degrading enzymes. Here, we characterize the biochemical properties of the symbiosis-induced polygalacturonase LbGH28A from the ectomycorrhizal fungus Laccaria bicolor. The transcriptional regulation of LbGH28A was measured by qPCR. The biological relevance of LbGH28A was confirmed by generating RNAi-silenced LbGH28A mutants. We localized the LbGH28A protein by immunofluorescence confocal and immunogold cytochemical microscopy in poplar ectomycorrhizal roots. qPCR confirmed the induced expression of LbGH28A during ectomycorrhiza formation. L. bicolor RNAi mutants have a lower ability to establish ectomycorrhiza confirming the key role of this enzyme in symbiosis. The purified recombinant LbGH28A has its highest activity towards pectin and polygalacturonic acid. In situ localization of LbGH28A indicates that this endopolygalacturonase is located in both fungal and plant cell walls at the symbiotic hyphal front. The present findings suggest that the symbiosis-induced pectinase LbGH28A is involved in the Hartig net formation and is an important determinant for successful symbiotic colonization.