Periodic Reporting for period 4 - PRO2EUK (Exploring Asgard archaea to illuminate the prokaryote-to-eukaryote transition)
Reporting period: 2023-12-01 to 2024-05-31
Cellular life on our planet is currently classified into three domains: Bacteria, Archaea and Eukarya ("eukaryotes"). Whereas the cytological properties of Bacteria and Archaea are relatively simple, eukaryotes are characterized by a high degree of cellular complexity. The origin of the eukaryotic cell is regarded as one of the major evolutionary innovations in the history of life on our planet. Yet, the emergence of the complex and compartmentalized nature of eukaryotic cells represents a major conundrum in modern biology. The central aim of the present ERC project "PRO2EUK" is to provide new insights in the enigmatic origin and evolution of the eukaryotic cell. The central question that is addressed in this ERC project is "How did complex (eukaryotic) life evolve?". In a broader context, this project is relevant to uncovering our own origins, as humans, together with other complex cellular life forms such as animals, plants, fungi etc, constitute the eukaryotic domain of life. As such, by understanding how eukaryotic life evolved, this will also provide insights into the very beginning of the lineage in the tree fo life that eventually gave rise to the emergence of us, humans. So, even though the central question is in essence a very fundamental one, it bears relevance to obtaining answers about our own origins - a recurring and hotly debated topic throughout human history.
The overall objectives of the project are:
• To explore the diversity of the Asgard archaeal superphylum at the genomic level using a variety of cultivation-independent approaches.
• To resolve the phylogenetic position of eukaryotes in the Tree of Life with respect to the improved taxonomic sampling of Asgard archaeal lineages and the use of state-of-the-art phylogenomics approaches.
• To use state-of-the-art ancestral genome reconstruction algorithms to determine the genome content of the Asgard archaeal ancestor of eukaryotes, providing insights into its cell biology and physiology in light of eukaryogenesis.
• To perform innovative in situ culturing techniques to identify potential syntrophic interaction networks that comprise Asgard archaea.
• To obtain high-resolution images of Asgard archaeal cells to provide insight in basic cell biological features of these archaea, and to infer cellular characteristics of the last common ancestor of archaea and eukaryotes.
The overall objectives of the project are:
• To explore the diversity of the Asgard archaeal superphylum at the genomic level using a variety of cultivation-independent approaches.
• To resolve the phylogenetic position of eukaryotes in the Tree of Life with respect to the improved taxonomic sampling of Asgard archaeal lineages and the use of state-of-the-art phylogenomics approaches.
• To use state-of-the-art ancestral genome reconstruction algorithms to determine the genome content of the Asgard archaeal ancestor of eukaryotes, providing insights into its cell biology and physiology in light of eukaryogenesis.
• To perform innovative in situ culturing techniques to identify potential syntrophic interaction networks that comprise Asgard archaea.
• To obtain high-resolution images of Asgard archaeal cells to provide insight in basic cell biological features of these archaea, and to infer cellular characteristics of the last common ancestor of archaea and eukaryotes.
A main objective of the proposal was to Genomic explore the diversity of the Asgard archaea: As part of WP1 we obtained several samples of mostly anaerobic sediments that were screened using 16S rRNA primers to capture the microbial diversity that was present. We found that marine sediments from Aarhus Bay generally contained the highest diversity and relative abundances, and we decided to focus on these sediments for follow up analyses. This choice was also practical, as it was relatively easy to obtain samples via our collaborators at Aarhus University (Denmark).
Next, we used genome-resolved metagenomics of Asgard archaeain order to study their genomic content, and to perform phylogenomic analyses. We chose a number of samples that were enriched in Asgard archaea for metagenomic sequencing (WP2). Using deep sequencing with both short and long read technologies, we managed to generate several high quality draft Asgard archaeal genomes (MAGs) that were included in follow up phylogenomic analyses. Part of this work was published in scientific journals (e.g. Eme et al 2023, Nature; Tamarit et al 2022, Nature Microbiology).
Subsequently, we used the generated genomic data of Asgard archaea, supplemented with publicly available data, to build a phylogenomic dataset that was analysed to determine the phylogenetic position of eukaryotes in the tree of life. Using a variety of phylogenomic approaches we established that eukaryotes are embedded within the Heimdallarchaeia, as a sister group of the Hodarchaeales. Furthermore, we used ancestral reconstruction analyses to trace the evolution of gene content across the Asgard archaea, including the lineage leading towards the eukaryotes. We found that the Asgard archaeal ancestor of eukaryotes (LAECA, see DoA) likely was an mesophilic, anaerobic heterotroph, and that the evolution of its gene content potentially showed traces of eukaryotic like-processes (e.g. elevated gene duplication rates). The results of these analyses were published in the scientific journal Nature (Eme et al, 2023, Nature).
Next, we aimed to establish an in situ cultivation approach to detect potential syntrophic interactions between Asgard archaea and their anticipated syntrophic partners. Rather then the original plan, in which we planned to use hydrogel beads and which turned out challenging, we decided to focus on developing a microfluidics-based cultivation approach, alongside more traditional enrichment cultivation techniques. The microfluidics-based cultivation approach was established up to a proof of principle stage, and will be further optimized, after which it will be published. Using traditional enrichment cultivation techniques, we managed to obtain several enrichments of Asgard archaeal lineages. The established cultivation approach also resulted in the enrichment of a novel methanogenic lineage outside of the Euryarchaeota (Wu et al, 2024, Nature).
Finally, we managed to obtain preliminary high-resolution images of Asgard archaeal cells using super-resolution microscopy and cryo-EM tomography techniques to provide insight in basic cell biological features of these archaea, and to infer cellular characteristics of the last common ancestor of archaea and eukaryotes. We are sill in the process of optimising these analyses, which are extremely challenging due to the slow growth speed of these organisms. Yet, we hope to conclude and publish the results of these analyses within the coming years.
Next, we used genome-resolved metagenomics of Asgard archaeain order to study their genomic content, and to perform phylogenomic analyses. We chose a number of samples that were enriched in Asgard archaea for metagenomic sequencing (WP2). Using deep sequencing with both short and long read technologies, we managed to generate several high quality draft Asgard archaeal genomes (MAGs) that were included in follow up phylogenomic analyses. Part of this work was published in scientific journals (e.g. Eme et al 2023, Nature; Tamarit et al 2022, Nature Microbiology).
Subsequently, we used the generated genomic data of Asgard archaea, supplemented with publicly available data, to build a phylogenomic dataset that was analysed to determine the phylogenetic position of eukaryotes in the tree of life. Using a variety of phylogenomic approaches we established that eukaryotes are embedded within the Heimdallarchaeia, as a sister group of the Hodarchaeales. Furthermore, we used ancestral reconstruction analyses to trace the evolution of gene content across the Asgard archaea, including the lineage leading towards the eukaryotes. We found that the Asgard archaeal ancestor of eukaryotes (LAECA, see DoA) likely was an mesophilic, anaerobic heterotroph, and that the evolution of its gene content potentially showed traces of eukaryotic like-processes (e.g. elevated gene duplication rates). The results of these analyses were published in the scientific journal Nature (Eme et al, 2023, Nature).
Next, we aimed to establish an in situ cultivation approach to detect potential syntrophic interactions between Asgard archaea and their anticipated syntrophic partners. Rather then the original plan, in which we planned to use hydrogel beads and which turned out challenging, we decided to focus on developing a microfluidics-based cultivation approach, alongside more traditional enrichment cultivation techniques. The microfluidics-based cultivation approach was established up to a proof of principle stage, and will be further optimized, after which it will be published. Using traditional enrichment cultivation techniques, we managed to obtain several enrichments of Asgard archaeal lineages. The established cultivation approach also resulted in the enrichment of a novel methanogenic lineage outside of the Euryarchaeota (Wu et al, 2024, Nature).
Finally, we managed to obtain preliminary high-resolution images of Asgard archaeal cells using super-resolution microscopy and cryo-EM tomography techniques to provide insight in basic cell biological features of these archaea, and to infer cellular characteristics of the last common ancestor of archaea and eukaryotes. We are sill in the process of optimising these analyses, which are extremely challenging due to the slow growth speed of these organisms. Yet, we hope to conclude and publish the results of these analyses within the coming years.
Pinpointing the position of eukaryotes in the tree of life (achievement in WP2) using advanced phylogenomics approaches can be considered an achievement significantly beyond the state of the art. Phylogenomic analyses aiming to resolve ancient evolutionary events, such as the origin of eukaryotes, are extremely challenging. By expanding the genomic sampling of Asgard archaeal genome space, in combination with development of a new phylogenomics pipeline, helped us to determine a new position of eukaryotes in the tree of life. Still, as we are in the progress of expanding these these analyses with a yet further expanded dataset of Asgard archaeal genome data, the obtained results might actually be fune-tuned further, providing additional insights into the elusive origin of eukaryotes.
Beyond genomics and phylogeneomics, we managed to obtain the first high resolution images of new groups of Asgard archaeal cells that seemingly point at a certain level of cellular complexity. These images were however of limited quality and number, which precludes us from making inferences about the cellular ultrastructure of Asgard archaea, and, by inference, of the last common ancestor of (Asgard) archaea and eukaryotes (LAECA). In case we would be able to obtain high resolution images of Asgard archaeal cells, this could potentially provide new insights about the cellular nature of LAECA, and in particular, about whether this ancestor already displayed a certain level of cellular complexity, or if this evolved later during eukaryotic evolution.
Beyond genomics and phylogeneomics, we managed to obtain the first high resolution images of new groups of Asgard archaeal cells that seemingly point at a certain level of cellular complexity. These images were however of limited quality and number, which precludes us from making inferences about the cellular ultrastructure of Asgard archaea, and, by inference, of the last common ancestor of (Asgard) archaea and eukaryotes (LAECA). In case we would be able to obtain high resolution images of Asgard archaeal cells, this could potentially provide new insights about the cellular nature of LAECA, and in particular, about whether this ancestor already displayed a certain level of cellular complexity, or if this evolved later during eukaryotic evolution.