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Report

Teaser, summary, work performed and final results

Periodic Reporting for period 2 - HeteroDynamic (Evolutionary Stability of Ubiquitous Root Symbiosis)

Teaser

Summary

Virtually all terrestrial plants depend on symbiotic interactions with fungi. Arbuscular mycorrhizal (AM) fungi evolved over 450 million years ago, are obligate biotrophs and cannot complete their life cycle without obtaining carbon from host roots. Mediating nutrient uptake and sequestering carbon in soil this symbiosis lie at the core of all terrestrial ecosystems. Plants on the other hand are facultative mycotrophs but under natural conditions all host roots are colonized as a result of multiple beneficial effects of AM fungi. In the symbiosis, both plants and fungi are promiscuous, forming interactions across individuals and species. In the absence of host-symbiont specificity and given their inability to discriminate among partners prior to interaction, evolutionary theory predicts that â€œfree ridersâ€ would evolve and spread. Yet AM fungi remain evolutionary and ecologically successful. I propose that this is thanks to their unique genomic organization, a temporally dynamic heterokaryosis.

In the HeteroDynamic research program we address the evolutionary stability of AM symbiosis, and base our work on the conceptual hypotehsis that the symbiosis is stable thanks to the genomic organization of the fungal partner, a temporally dynamic heterokaryosis. This hypothesis is based on the assumptions that heterokaryosis allows for functional variation over time - and my team combines bioinformatic and experimental work to test this.

Our research is not only important for better understanding evolutionary dynamics of ancient mutualistic interactions between fungi and plants but are potentially important for future agriculture including the use of AM inoculum, optimized rotations and breeding for beneficial AM fungi. While negative feedback between fungi and host plants is important in maintaining high biodiversity in natural ecosystems its role in agriculture has not yet been tested and our studies build the foundation for such studies. To take advantage of growth promoting AM fungal communities in sustainable intensification of agriculture, systems need to be developed where crop rotations are tested for their ability to promote continually beneficial communities.

Work performed

Since the start of the HeteroDynamic program we have generated extensive sequence datasets and assembled these to build the foundation to test our central hypothesis.

Firstly we developed a novel single nucleus genomics method for sequencing and assembly of AM fungal genomes from single nuclei. Reads from individual nuclei can then be mapped back to the assembly to evaluate sequence variance among nuclei from individual strains and even from within single spores. The physical sorting of nuclei using FACS had been tested for one species prior to the onset of the HeteoDynamic research program. During the project the method has been developed to accommodate a range of species and a custom-made assembly workflow has been developed to handle the generated data. We are currently in the process of publishing the method. An added value of this method is that it allows us to assemble genomes from a broad range of AM fungi that could previously not be sequenced due to challenges obtaining sufficient amount and purity of biological material from most AM fungi.

Secondly we test for functional heterokaryosis, by simultaneously exposing fungal strains to different hosts. Plant and fungal activity in different host â€“ symbiont combinations will be tested using RNA and mapping differentially expressed genes onto single nuclei reads for the sequenced fungal strains. This will allow us to link host specific expression to potential nucleotype variation.

Finally, we address temporal variation in nucleotype composition by performing dual host segregation experiments where we will test for fungal and plant fitness in interactions involving fungal strains adapted to different host. The central hypothesis here is that the nucleotypes that are most fit on a host will become more abundant on that host leading to increased fitness of the adapted fungal strain and decreased fitness of the plant host. Quantification of carbon and phosphorus trade in symbiotic interactions involving adapted and non adapted fungal strains will be performed.

Growth experimental systems for the two later have been developed during the first part of the project and we now have enough experience and know how to perform these experiments.

Final results

Our single nuclei sequencing method takes the project beyond the state of the art and by the end of this research program we will be able to make important contributions to the field of AM genomics and evolution.