In vitro resynthesis of lichenization reveals the genetic background of symbiosis-specific fungal-algal interaction in Usnea hakonensis.

Abstract

Background Symbiosis is central to ecosystems and has been an important driving force of the diversity of life. Close and long-term interactions are known to develop cooperative molecular mechanisms between the symbiotic partners and have often given them new functions as symbiotic entities. In lichen symbiosis, mutualistic relationships between lichen-forming fungi and algae and/or cyanobacteria produce unique features that make lichens adaptive to a wide range of environments. Although the morphological, physiological, and ecological uniqueness of lichens has been described for more than a century, the genetic mechanisms underlying this symbiosis are still poorly known.Results This study investigated the fungal-algal interaction specific to the lichen symbiosis using Usnea hakonensis as a model system. The whole genome of U. hakonensis, the fungal partner, was sequenced by using a culture isolated from a natural lichen thallus. Isolated cultures of the fungal and the algal partners were co-cultured in vitro for three months, and thalli were successfully resynthesized as visible protrusions. Transcriptomes of resynthesized and natural thalli (symbiotic states) were compared to that of isolated cultures (non-symbiotic state). Sets of fungal and algal genes up-regulated in both symbiotic states were identified as symbiosis-related genes.Conclusion From predicted functions of these genes, we identified genetic association with two key features fundamental to the symbiotic lifestyle in lichens. The first is establishment of a fungal symbiotic interface: (a) modification of cell walls at fungal-algal contact sites; and (b) production of a hydrophobic layer that ensheaths fungal and algal cells;. The second is symbiosis-specific nutrient flow: (a) the algal supply of photosynthetic product to the fungus; and (b) the fungal supply of phosphorous and nitrogen compounds to the alga. Since both features are widespread among lichens, our result may indicate important facets of the genetic basis of the lichen symbiosis.

Lichen algae: the photosynthetic partners in lichen symbioses

“…Whether any substance is transferred from fungus to alga in exchange has yet to be demonstrated. At least some genes relevant to such metabolic transfers appear to be differentially expressed in symbiosis (Kono et al 2020). Certainly, there has been speculation that the fungal partner might apportion carbohydrate, nitrogen, or other substances back to the algal symbiont to regulate its growth (Ahmadjian 1995) in coordination with that of the mycobiont (Greenhalgh & Anglesea 1979;Hill 1985Hill , 1989Honegger 1987).…”

Phylogenomics reveals the evolutionary origins of lichenization in chlorophyte algae

“…To further identify among these features those directly associated with the ability to lichenize, we cross-referenced the genes containing the discriminating IPR and PFAM terms with the genes found up-regulated in the LFA species Trebouxia sp . TZW2008 ( SI Appendix , Table S6) in association with its fungal symbiont Usnea hakonensis (19). After this cross-referencing, we retained 35 IPR mainly present in LFA, and 115 IPR significantly expanded in LFA ( SI Appendix , Table S4).…”

“…Along with glycosyl hydrolases, we also identified the expansions of the short-chain dehydrogenase gene family putatively involved in the biosynthesis and excretion of ribitol. Ribitol has been demonstrated to impact fungal growth in various lichens and is thought to act as a signal to initiate lichen formation (31), and its biosynthesis has been found regulated in the lichens Cladonia grayi and Usnea hakonensis (18, 19).…”

“…In order to increase our statistical power by crossing deregulated genes in symbiotic context we used the transcriptomic data from Trebouxia sp TZW2008 (19) with results of our contraction/expansion analysis. To do so, the raw reads were downloaded ( SI Appendix , Table S1) and submitted to the nf-core/rnaseq v3.4 (95) workflow in nextflow v21.04 (96) using -profile debug,genotoul --skip_qc --aligner star_salmon options.…”

“…To empower the statistical approaches, transcriptomic data from Trebouxia sp. (TZW2008 isolate, 21 ) was used to analyze significative IPR/orthogroups that also corresponds to a DEG in Trebouxia sp. TZW2008.…”