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Description
Seabed hot vents are one of the most poorly understood habitats on Earth. They provide a broad range for diverse and extreme habitats with steep temperature and pH-gradients where diverse extremophilic microorganisms thrive. It is now well-established that the life in these extreme and inhospitable environments, is fueled by chemosynthesis, rather than photosynthesis, and that these habitats are our window into the first lifeforms to have emerged on our planet. Importantly, they also represent a biosphere significant for bioprospecting and biotechnology. Unfortunately, still, the vast majority of the microbial functional and biochemical diversity operating in these habitats is hidden in uncultured and yet-uncharacterized lineages. Therefore, starting at the unique biodiversity of the vents found under Norwegian waters at the Arctic Mid-Ocean Ridge, INDEPTH aims to decipher the metabolic traits and biochemical/enzymological content of this hidden reservoir. Through Polish and Norwegian complementarity of the research experience, technical expertise, knowledge and resources, and distribution, we will combine and leverage our expertise on extremophilic enzymes in Gdansk and Poznan, adaptations of extremophiles in Warsaw and ecology and bioprospecting of hot vents in Bergen. We will integrate and develop our competences in five work packages, focusing on bioinformatics-based metabolic predictions and phylogeny (WP1), adaptations (WP2), and structure/function analyses of enzymes (WP3, WP4, WP5). Fundamentally important results can be expected with substantial impact on our understanding of the ecology of microbial metabolism with the focus on carbon biogeochemical cycling, its evolution and the underlying molecular adaptations to life under the extreme conditions. The project workflow has the potential to unleash an enormous resource and improve the knowledge of enzymes operating under extreme conditions, similar to those that are of interest for industrial applications.
Summary of project results
The deep sea covers over half of the Earth''s surface area. With an average depth of almost 4000 meters, the deep ocean basins are the most poorly explored places on Earth. It is estimated that the deep sea contains 80% of Earth''s biosphere and represents the biggest living system on Earth, distinctive in its unparalleled complexity, although massively integrated at all levels of ecological organization. This enormous biosphere is often considered life''s last frontier, characterized by lack of light, high pressure, limited supply of nutrients, and temperatures ranging from low (2-4°C) in the majority of habitats to extremely high (>100°C) in the seabed hot hydrothermal vents. This biosphere''s dimension, diversity, and functioning are almost beyond comprehension. It is well-established that life in these extreme and hostile environments is fuelled by chemosynthesis - rather than photosynthesis –that they are windows into the first lifeforms to have emerged on our planet and represent a biosphere significant for bioprospecting and biotechnology. Unfortunately, most of these habitats'' microbial functional and biochemical diversity is still hidden in uncultured yet uncharacterized lineages. Therefore, starting at the unique biodiversity of the vents found under Norwegian waters at the Arctic Mid-Ocean Ridge, INDEPTH''s key objective was to decipher this hidden reservoir''s metabolic traits and biochemical/enzymological content. Through Polish and Norwegian complementarity and distribution, we combined expertise on extremophilic enzymes in Gdansk (University of Gdansk) and Poznan (Institute of Bioorganic Chemistry), adaptations of extremophiles in Warsaw (University of Warsaw), and ecology and bioprospecting of hot vents in Bergen (University of Bergen, Norway). We integrated and developed our competence in the following areas: (i) bioinformatics-based metabolic predictions and phylogeny, (ii) adaptations, and (iii) structure/function analyses of enzymes.
The efforts in reconstructing and analyzing microbial genomes from the Arctic Mid-Ocean Ridges hydrothermal vent fields have led to significant discoveries within heterotrophic lineages involved in cycling organic carbon in marine environments. In particular, in the archaeal Korarchaeia, the potential for homoacetogenesis via the Wood-Ljungdahl pathway (WLP) was elucidated, confirming previous hypotheses that the WLP may have evolved independently from methanogenic metabolism in Archaea. In summary, we show the genetic link that deep-branching Korarchaeia can couple fermentation to acetate production and perform homoacetogenesis via the WLP. This metabolic feature is a major finding in the class Korarchaeia from extreme deep-sea habitats. In addition, the use of advanced bioinformatics allowed us to identify unique adaptation strategies among microbes inhabiting deep-sea floors concerning resistance to heavy metals, high temperature, and metabolism of C1-type compounds.
Another research activity was focused on developing a bioinformatic tool for searching proteins with novel functions. The INDEPTH biodiscovery pipeline is based on a unique algorithm to detect protein domain profiles characteristic of certain groups of enzymes. The targets selected using this innovative approach included DNA polymerases, nucleases, single-stranded DNA binding proteins, DNA helicases, laccases, beta-galactosidases, chitinases, alkaline phosphatases, pectinases, proteinases, and petases. Biochemical and biophysical methods allowed us to prove their extreme thermal stability, robust activity, and ability to operate under extreme conditions, similar to those of interest for industrial applications.
In addition to research activities, the INDEPTH project focused on transferring knowledge about the exploitation of complex microbial communities into teaching and training. Indeed, we firmly believe that INDEPTH’s long-lasting legacy is teaching and training the next generation of scientists, from Bachelor and Master students to postgraduates and early-career researchers.
Better understanding of the biodiversity of microbial communities thriving in extreme deep-sea habitats. The analysis of microbial genomes from the Arctic Mid-Ocean Ridges hydrothermal vent fields has led to discoveries within heterotrophic lineages involved in cycling organic carbon in marine environments. In the archaeal Korarchaeia, the potential for homoacetogenesis via the Wood-Ljungdahl pathway was elucidated, confirming hypotheses that this pathway may have evolved independently from methanogenic metabolism in Archaea. It was shown by University of Bergen team that deep-branching Korarchaeia can couple fermentation to acetate production and perform homoacetogenesis. This metabolic feature is a major finding in the class Korarchaeia from extreme deep-sea habitats.
Development of strategies for bioprospecting of microbial metagenomes. The INDEPTH biodiscovery pipeline developed by the University of Warsaw research team is based on detecting protein domain profiles characteristic of certain groups of enzymes. A domain-based filtering approach, built on Hidden Markov Models, was employed to identify proteins potentially possessing relevant enzymatic function. The consortium''s efforts to analyze the properties of target enzymes were led by the University of Gdansk. It resulted in characterization of >30 novel biocatalysts.
Structural features of novel enzymes. Three novel crystal structures have been obtained by the Institute of Bioorganic Chemistry group, two at high- resolution (S1/P1 nuclease and Thermus phage Tt72 endolysin) and one at low resolution (metagenomic PolA-type DNA polymerase).
Analysis of adaptation strategies among microbes inhabiting deep-sea floor. The following features were analysed by University of Warsaw team: (i) metal resistance, (ii) adaptation to extreme thermal conditions, and (iii) metabolism of C1-type compounds. The analysis revealed the presence of genes conferring resistance to arsen, chrom, copper, cobalt, zinc, cadmium, mercury, and nickel. Among the determinants responsible for cold tolerance, the most common were genes coding for cold shock proteins. On the other hand, in the group of heat shock genes, the most numerous were genes whose products interacted with DnaJ, DnaK, and GrpE. Analysis of the distribution of genes related to the metabolism of C1-type compounds revealed the presence of metabolic pathways related to methanogenesis, methanotrophy, and methylotrophy in the studied environments.
Summary of bilateral results
The project strengthened bilateral cooperation between beneficiary and donor state entities. Collaboration between project partners from Poland and Norway allowed knowledge transfer and provided significant educational opportunities to young scientists associated with participating institutions known for their scientific excellence in basic research. Through this project it was possible to master teamwork, share best practices, exchange information and know-how between research groups, thus contributing to European cohesion objectives by network activities. Interaction with peers by attending international conferences created a platform to exchange ideas to solve scientific problems. The project provided excellent opportunities in basic research for a number of Polish and Norwegian early-career researchers, M.Sc. students, Ph.D. students and postdoctoral fellows who were working together to achieve goals outlined in the project proposal. A number of them were working directly in the project through practical training in partners'' laboratories. The partners applied for an EU Twinning grant (HORIZON WIDERA 2023) with the proposal VISIONARIES, which passed the evaluation threshold but did not receive funding.