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The Asgard archaeal origins of Arf family GTPases involved in eukaryotic organelle dynamics

Nature Microbiology volume 10, pages 495–508 (2025)Cite this article

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Abstract

The evolution of eukaryotes is a fundamental event in the history of life. The closest prokaryotic lineage to eukaryotes, the Asgardarchaeota, encode proteins previously found only in eukaryotes, providing insight into their archaeal ancestor. Eukaryotic cells are characterized by endomembrane organelles, and the Arf family GTPases regulate organelle dynamics by recruiting effector proteins to membranes upon activation. The Arf family is ubiquitous among eukaryotes, but its origins remain elusive. Here we report a group of prokaryotic GTPases, the ArfRs, which are widely present in Asgardarchaeota. Phylogenetic analyses reveal that eukaryotic Arf family proteins arose from the ArfR group. Expression of representative Asgardarchaeota ArfR proteins in yeast and X-ray crystallographic studies show that ArfR GTPases possess the mechanism of membrane binding and structural features unique to Arf family proteins. Our results indicate that Arf family GTPases originated in the archaeal ancestor of eukaryotes, consistent with aspects of the endomembrane system evolving early in eukaryogenesis.

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Fig. 1: Phylogeny of the Ras superfamily.
Fig. 2: Distribution of the major subgroups of the Ras superfamily across different lineages of Asgardarchaeota.
Fig. 3: Localization of asgardarchaeote ArfR proteins in yeast cells.
Fig. 4: The N-terminal AH of asgardarchaeote ArfR proteins is required for membrane localization in yeast cells.
Fig. 5: GDP/GTP structural cycle of HodArfR1.
Fig. 6: Structural comparison of the GDP-bound form of asgardarchaeote and eukaryotic Arf family proteins.

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Data availability

All DNA and protein sequences analysed in this study come from public resources specified in supplementary tables and figures. Phylogenetic trees in the NEWICK format and the underlying sequence alignments are available via figshare at https://doi.org/10.6084/m9.figshare.26830264 (ref. 96). The diffraction data and 3D structures reported in this study are available at the World Wide Protein Data Bank (https://www.wwpdb.org/) under PDB accession numbers 8OUK, 8OUL, 8OUM and 8OUN. The GerdArfR1 and HodArfR1 E. coli and S. cerevisiae expression plasmids are available upon request from J.M. and C.L.J., respectively.

Code availability

The script used in the analysis reported in this paper is available at https://github.com/Losssina/ScrollSaw.git

References

  1. Donoghue, P. C. J. et al. Defining eukaryotes to dissect eukaryogenesis. Curr. Biol. 33, R919–R929 (2023).

    CAS  PubMed  Google Scholar 

  2. Spang, A. et al. Proposal of the reverse flow model for the origin of the eukaryotic cell based on comparative analyses of Asgard archaeal metabolism. Nat. Microbiol. 4, 1138–1148 (2019).

    CAS  PubMed  Google Scholar 

  3. López-García, P. & Moreira, D. The symbiotic origin of the eukaryotic cell. C. R. Biol. 346, 55–73 (2023).

    PubMed  Google Scholar 

  4. Baum, D. A. & Baum, B. An inside-out origin for the eukaryotic cell. BMC Biol. 12, 75 (2014).

    Google Scholar 

  5. Vosseberg, J. et al. The emerging view on the origin and early evolution of eukaryotic cells. Nature 633, 295–305 (2024).

    CAS  PubMed  Google Scholar 

  6. Spang, A. et al. Complex archaea that bridge the gap between prokaryotes and eukaryotes. Nature 521, 173–179 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Zaremba-Niedzwiedzka, K. et al. Asgard archaea illuminate the origin of eukaryotic cellular complexity. Nature 541, 353–358 (2017).

    CAS  PubMed  Google Scholar 

  8. Eme, L. et al. Inference and reconstruction of the heimdallarchaeial ancestry of eukaryotes. Nature 618, 992–999 (2023).

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Liu, Y. et al. Expanded diversity of Asgard archaea and their relationships with eukaryotes. Nature 593, 553–557 (2021).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Hatano, T. et al. Asgard archaea shed light on the evolutionary origins of the eukaryotic ubiquitin-ESCRT machinery. Nat. Commun. 13, 3398 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Neveu, E., Khalifeh, D., Salamin, N. & Fasshauer, D. Prototypic SNARE proteins are encoded in the genomes of Heimdallarchaeota, potentially bridging the gap between the prokaryotes and eukaryotes. Curr. Biol. 30, 2468–2480.e5 (2020).

    CAS  PubMed  Google Scholar 

  12. Akıl, C. & Robinson, R. C. Genomes of Asgard archaea encode profilins that regulate actin. Nature 562, 439–443 (2018).

    PubMed  Google Scholar 

  13. Akıl, C. et al. Structural and biochemical evidence for the emergence of a calcium-regulated actin cytoskeleton prior to eukaryogenesis. Commun. Biol. 5, 890 (2022).

    PubMed  PubMed Central  Google Scholar 

  14. Rodrigues-Oliveira, T. et al. Actin cytoskeleton and complex cell architecture in an Asgard archaeon. Nature 613, 332–339 (2023).

    CAS  PubMed  Google Scholar 

  15. Survery, S. et al. Heimdallarchaea encodes profilin with eukaryotic-like actin regulation and polyproline binding. Commun. Biol. 4, 1024 (2021).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Akıl, C. et al. Structure and dynamics of Odinarchaeota tubulin and the implications for eukaryotic microtubule evolution. Sci. Adv. 8, eabm2225 (2022).

    PubMed  PubMed Central  Google Scholar 

  17. Donaldson, J. G. & Jackson, C. L. ARF family G proteins and their regulators: roles in membrane transport, development and disease. Nat. Rev. Mol. Cell Biol. 12, 362–375 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Li, F.-L. & Guan, K.-L. The Arf family GTPases: regulation of vesicle biogenesis and beyond. Bioessays 45, 2200214 (2023).

    CAS  Google Scholar 

  19. Jackson, C. L., Ménétrey, J., Sivia, M., Dacks, J. B. & Eliáš, M. An evolutionary perspective on Arf family GTPases. Curr. Opin. Cell Biol. 85, 102268 (2023).

    CAS  PubMed  Google Scholar 

  20. Sztul, E. et al. ARF GTPases and their GEFs and GAPs: concepts and challenges. Mol. Biol. Cell 30, 1249–1271 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Antonny, B., Beraud-Dufour, S., Chardin, P. & Chabre, M. N-terminal hydrophobic residues of the G-protein ADP-ribosylation factor-1 insert into membrane phospholipids upon GDP to GTP exchange. Biochemistry 36, 4675–4684 (1997).

    CAS  PubMed  Google Scholar 

  22. Amor, J. C., Harrison, D. H., Kahn, R. A. & Ringe, D. Structure of the human ADP-ribosylation factor 1 complexed with GDP. Nature 372, 704–708 (1994).

    CAS  PubMed  Google Scholar 

  23. Gillingham, A. K. & Munro, S. The small G proteins of the Arf family and their regulators. Annu. Rev. Cell Dev. Biol. 23, 579–611 (2007).

    CAS  PubMed  Google Scholar 

  24. Pasqualato, S., Renault, L. & Cherfils, J. Arf, Arl, Arp and Sar proteins: a family of GTP-binding proteins with a structural device for ‘front–back’ communication. EMBO Rep. 3, 1035–1041 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Greasley, S. E. et al. The structure of rat ADP-ribosylation factor-1 (ARF-1) complexed to GDP determined from two different crystal forms. Nat. Struct. Biol. 2, 797–806 (1995).

    CAS  PubMed  Google Scholar 

  26. Ménétrey, J., Macia, E., Pasqualato, S., Franco, M. & Cherfils, J. Structure of Arf6–GDP suggests a basis for guanine nucleotide exchange factors specificity. Nat. Struct. Biol. 7, 466–469 (2000).

    PubMed  Google Scholar 

  27. Goldberg, J. Structural basis for activation of ARF GTPase: mechanisms of guanine nucleotide exchange and GTP–myristoyl switching. Cell 95, 237–248 (1998).

    CAS  PubMed  Google Scholar 

  28. Pasqualato, S., Ménétrey, J., Franco, M. & Cherfils, J. The structural GDP/GTP cycle of human Arf6. EMBO Rep. 2, 234–238 (2001).

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Bi, X., Corpina, R. A. & Goldberg, J. Structure of the Sec23/24–Sar1 pre-budding complex of the COPII vesicle coat. Nature 419, 271–277 (2002).

    CAS  PubMed  Google Scholar 

  30. Panic, B., Perisic, O., Veprintsev, D. B., Williams, R. L. & Munro, S. Structural basis for Arl1-dependent targeting of homodimeric GRIP domains to the golgi apparatus. Mol. Cell 12, 863–874 (2003).

    CAS  PubMed  Google Scholar 

  31. Mourão, A., Nager, A. R., Nachury, M. V. & Lorentzen, E. Structural basis for membrane targeting of the BBSome by ARL6. Nat. Struct. Mol. Biol. 21, 1035–1041 (2014).

    PubMed  PubMed Central  Google Scholar 

  32. Lokaj, M. et al. The interaction of CCDC104/BARTL1 with Arl3 and implications for ciliary function. Structure 23, 2122–2132 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Surkont, J. & Pereira-Leal, J. B. Are there Rab GTPases in archaea? Mol. Biol. Evol. 33, 1833–1842 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Tran, L. T., Akıl, C., Senju, Y. & Robinson, R. C. The eukaryotic-like characteristics of small GTPase, roadblock and TRAPPC3 proteins from Asgard archaea. Commun. Biol. 7, 273 (2024).

    CAS  PubMed  PubMed Central  Google Scholar 

  35. Homma, Y., Hiragi, S. & Fukuda, M. Rab family of small GTPases: an updated view on their regulation and functions. FEBS J. 288, 36–55 (2021).

    CAS  PubMed  Google Scholar 

  36. Klinger, C. M., Spang, A., Dacks, J. B. & Ettema, T. J. G. Tracing the archaeal origins of eukaryotic membrane-trafficking system building blocks. Mol. Biol. Evol. 33, 1528–1541 (2016).

    CAS  PubMed  Google Scholar 

  37. Vargová, R. et al. A eukaryote-wide perspective on the diversity and evolution of the ARF GTPase protein family. Genome Biol. Evol. 13, evab157 (2021).

    PubMed  PubMed Central  Google Scholar 

  38. Parks, D. H. et al. Recovery of nearly 8,000 metagenome-assembled genomes substantially expands the tree of life. Nat. Microbiol. 2, 1533–1542 (2017).

    CAS  PubMed  Google Scholar 

  39. Jadhav, B., Wild, K., Pool, M. R. & Sinning, I. Structure and switch cycle of SRβ as ancestral eukaryotic GTPase associated with secretory membranes. Structure 23, 1838–1847 (2015).

    CAS  PubMed  Google Scholar 

  40. Bigay, J. & Antonny, B. Curvature, lipid packing, and electrostatics of membrane organelles: defining cellular territories in determining specificity. Dev. Cell 23, 886–895 (2012).

    CAS  PubMed  Google Scholar 

  41. Vasudevan, C. et al. The distribution and translocation of the G protein ADP-ribosylation factor 1 in live cells is determined by its GTPase activity. J. Cell Sci. 111, 1277–1285 (1998).

    CAS  PubMed  Google Scholar 

  42. Imachi, H. et al. Isolation of an archaeon at the prokaryote–eukaryote interface. Nature 577, 519–525 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Garten, M. et al. Methyl-branched lipids promote the membrane adsorption of α-synuclein by enhancing shallow lipid-packing defects. Phys. Chem. Chem. Phys. 17, 15589–15597 (2015).

    CAS  PubMed  Google Scholar 

  44. Čopič, A. et al. A giant amphipathic helix from a perilipin that is adapted for coating lipid droplets. Nat. Commun. 9, 1332 (2018).

    PubMed  PubMed Central  Google Scholar 

  45. Fleming, J. R., Dawson, A. & Hunter, W. N. Crystal structure of Leishmania major ADP-ribosylation factor-like 1 and a classification of related GTPase family members in this kinetoplastid. Mol. Biochem. Parasitol. 174, 141–144 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Mills, J. et al. Extracellular vesicle formation in Euryarchaeota is driven by a small GTPase. Proc. Natl Acad. Sci. USA 121, e2311321121 (2024).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Valentin-Alvarado, L. E. et al. Complete genomes of Asgard archaea reveal diverse integrated and mobile genetic elements. Genome Res. 34, 1595–1609 (2024).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Akıl, C. et al. Insights into the evolution of regulated actin dynamics via characterization of primitive gelsolin/cofilin proteins from Asgard archaea. Proc. Natl Acad. Sci. USA 117, 19904–19913 (2020).

    PubMed  PubMed Central  Google Scholar 

  49. Lu, Z. et al. Coevolution of eukaryote-like Vps4 and ESCRT-III subunits in the Asgard archaea. mBio 11, e00417-20 (2020).

    PubMed  PubMed Central  Google Scholar 

  50. Nachmias, D. et al. Asgard ESCRT-III and VPS4 reveal conserved chromatin binding properties of the ESCRT machinery. ISME J. 17, 117–129 (2023).

    CAS  PubMed  Google Scholar 

  51. Altschul, S. F. et al. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25, 3389–3402 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Katoh, K. & Standley, D. M. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol. Biol. Evol. 30, 772–780 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  53. Hall, T. A. BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucl. Acids Symp. Ser. 41, 95–98 (1999).

    CAS  Google Scholar 

  54. Capella-Gutiérrez, S., Silla-Martínez, J. M. & Gabaldón, T. trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics 25, 1972–1973 (2009).

    PubMed  PubMed Central  Google Scholar 

  55. Nguyen, L.-T., Schmidt, H. A., von Haeseler, A. & Minh, B. Q. IQ-TREE: a fast and effective stochastic algorithm for estimating maximum-likelihood phylogenies. Mol. Biol. Evol. 32, 268–274 (2015).

    CAS  PubMed  Google Scholar 

  56. Guindon, S. et al. New algorithms and methods to estimate maximum-likelihood phylogenies: assessing the performance of PhyML 3.0. Syst. Biol. 59, 307–321 (2010).

    CAS  PubMed  Google Scholar 

  57. Minh, B. Q., Nguyen, M. A. T. & von Haeseler, A. Ultrafast approximation for phylogenetic bootstrap. Mol. Biol. Evol. 30, 1188–1195 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  58. Mirarab, S. et al. PASTA: ultra-large multiple sequence alignment for nucleotide and amino-acid sequences. J. Comput. Biol. 22, 377–386 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  59. Letunic, I. & Bork, P. Interactive tree of life (iTOL) v3: an online tool for the display and annotation of phylogenetic and other trees. Nucleic Acids Res. 44, W242–W245 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  60. Marchler-Bauer, A. & Bryant, S. H. CD-Search: protein domain annotations on the fly. Nucleic Acids Res. 32, W327–W331 (2004).

    CAS  PubMed  PubMed Central  Google Scholar 

  61. Lu, S. et al. CDD/SPARCLE: the conserved domain database in 2020. Nucleic Acids Res. 48, D265–D268 (2020).

    CAS  PubMed  Google Scholar 

  62. Leipe, D. D., Wolf, Y. I., Koonin, E. V. & Aravind, L. Classification and evolution of P-loop GTPases and related ATPases1. J. Mol. Biol. 317, 41–72 (2002).

    CAS  PubMed  Google Scholar 

  63. Seitz, K. W., Lazar, C. S., Hinrichs, K.-U., Teske, A. P. & Baker, B. J. Genomic reconstruction of a novel, deeply branched sediment archaeal phylum with pathways for acetogenesis and sulfur reduction. ISME J. 10, 1696–1705 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  64. Seitz, K. W. et al. Asgard archaea capable of anaerobic hydrocarbon cycling. Nat. Commun. 10, 1822 (2019).

    PubMed  PubMed Central  Google Scholar 

  65. Cai, M. et al. Diverse Asgard archaea including the novel phylum Gerdarchaeota participate in organic matter degradation. Sci. China Life Sci. 63, 886–897 (2020).

    CAS  PubMed  Google Scholar 

  66. Farag, I. F., Zhao, R. & Biddle, J. F. ‘Sifarchaeota,’ a novel Asgard phylum from Costa Rican sediment capable of polysaccharide degradation and anaerobic methylotrophy. Appl. Environ. Microbiol. 87, e02584-20 (2021).

  67. Sun, J. et al. Recoding of stop codons expands the metabolic potential of two novel Asgardarchaeota lineages. ISME Commun. 1, 30 (2021).

    PubMed  PubMed Central  Google Scholar 

  68. Xie, R. et al. Expanding Asgard members in the domain of Archaea sheds new light on the origin of eukaryotes. Sci. China Life Sci. 65, 818–829 (2022).

    CAS  PubMed  Google Scholar 

  69. Price, M. N., Dehal, P. S. & Arkin, A. P. FastTree 2 – approximately maximum-likelihood trees for large alignments. PLoS ONE 5, e9490 (2010).

    PubMed  PubMed Central  Google Scholar 

  70. Valentin-Alvarado, L. E. et al. Asgard archaea modulate potential methanogenesis substrates in wetland soil. Nat. Commun. 15, 6384 (2024).

    CAS  PubMed  PubMed Central  Google Scholar 

  71. Tamarit, D. et al. Description of Asgardarchaeum abyssi gen. nov. spec. nov., a novel species within the class Asgardarchaeia and phylum Asgardarchaeota in accordance with the SeqCode. Syst. Appl. Microbiol. 47, 126525 (2024).

    CAS  PubMed  Google Scholar 

  72. Parks, D. H. et al. GTDB: an ongoing census of bacterial and archaeal diversity through a phylogenetically consistent, rank normalized and complete genome-based taxonomy. Nucleic Acids Res. 50, D785–D794 (2022).

    CAS  PubMed  Google Scholar 

  73. Liu, R. et al. Metagenomic insights into Heimdallarchaeia clades from the deep-sea cold seep and hydrothermal vent. Environ. Microbiome 19, 43 (2024).

    CAS  PubMed  PubMed Central  Google Scholar 

  74. Appler, K. E. et al. Oxygen metabolism in descendants of the archaeal-eukaryotic ancestor. Preprint at bioRxiv https://doi.org/10.1101/2024.07.04.601786 (2024).

  75. Eliáš, M., Brighouse, A., Gabernet-Castello, C., Field, M. C. & Dacks, J. B. Sculpting the endomembrane system in deep time: high resolution phylogenetics of Rab GTPases. J. Cell Sci. 125, 2500–2508 (2012).

    PubMed  PubMed Central  Google Scholar 

  76. Vosseberg, J. et al. Timing the origin of eukaryotic cellular complexity with ancient duplications. Nat. Ecol. Evol. 5, 92–100 (2021).

    PubMed  Google Scholar 

  77. Schmidt, H. A., Strimmer, K., Vingron, M. & von Haeseler, A. TREE-PUZZLE: maximum likelihood phylogenetic analysis using quartets and parallel computing. Bioinformatics 18, 502–504 (2002).

    CAS  PubMed  Google Scholar 

  78. Goodstadt, L. & Ponting, C. P. CHROMA: consensus-based colouring of multiple alignments for publication. Bioinformatics 17, 845–846 (2001).

    CAS  PubMed  Google Scholar 

  79. Frickey, T. & Lupas, A. CLANS: a Java application for visualizing protein families based on pairwise similarity. Bioinformatics 20, 3702–3704 (2004).

    CAS  PubMed  Google Scholar 

  80. Gautier, R., Douguet, D., Antonny, B. & Drin, G. HELIQUEST: a web server to screen sequences with specific α-helical properties. Bioinformatics 24, 2101–2102 (2008).

    CAS  PubMed  Google Scholar 

  81. Pranke, I. M. et al. α-Synuclein and ALPS motifs are membrane curvature sensors whose contrasting chemistry mediates selective vesicle binding. J. Cell Biol. 194, 89–103 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  82. Huh, W.-K. et al. Global analysis of protein localization in budding yeast. Nature 425, 686–691 (2003).

    CAS  PubMed  Google Scholar 

  83. Kabsch, W. XDS. Acta Crystallogr. D 66, 125–132 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  84. Tickle, I. J. et al. STARANISO (Global Phasing, 2018).

  85. Vonrhein, C. et al. Data processing and analysis with the autoPROC toolbox. Acta Crystallogr. D 67, 293–302 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  86. McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  87. Kelley, L. A., Mezulis, S., Yates, C., Wass, M. N. & Sternberg, M. J. E. The Phyre2 web portal for protein modeling, prediction and analysis. Nat. Protoc. 10, 845–858 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  88. Zhu, W., Shenoy, A., Kundrotas, P. & Elofsson, A. Evaluation of AlphaFold-Multimer prediction on multi-chain protein complexes. Bioinformatics 39, btad424 (2023).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. Jumper, J. et al. Highly accurate protein structure prediction with AlphaFold. Nature 596, 583–589 (2021).

    CAS  PubMed  PubMed Central  Google Scholar 

  90. Mirdita, M. et al. ColabFold: making protein folding accessible to all. Nat. Methods 19, 679–682 (2022).

    CAS  PubMed  PubMed Central  Google Scholar 

  91. Terwilliger, T. C. et al. Iterative model building, structure refinement and density modification with the PHENIX AutoBuild wizard. Acta Crystallogr. D 64, 61–69 (2008).

    CAS  PubMed  Google Scholar 

  92. Perrakis, A., Morris, R. & Lamzin, V. S. Automated protein model building combined with iterative structure refinement. Nat. Struct. Biol. 6, 458–463 (1999).

    CAS  PubMed  Google Scholar 

  93. Bricogne G. et al. BUSTER Version 2.10.2 (Global Phasing, 2016).

  94. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D 60, 2126–2132 (2004).

    PubMed  Google Scholar 

  95. DeLano, W. L. The PyMOL Molecular Graphics System Version 1.2 (Schrödinger, 2002).

  96. Vargová, R. Arf family GTPases are present in Asgard archaea. figshare https://doi.org/10.6084/m9.figshare.26830264 (2024).

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Acknowledgements

We especially thank B. Gigant and V. Campanacci for help with X-ray data collection and scaling, C. Besse and M. Chenon for plasmid construction and T. Ettema for input in the initial stages of the project. We are very thankful to R. Kahn (Emory University), R. Karess and V. Campanacci for careful reading and constructive comments on the manuscript and D. Tamarit for valuable discussions regarding the ever-evolving asgardarchaeote classification. We are most grateful to the machine (SOLEIL synchrotron, Saint-Aubin, France) and beamline groups (PROXIMA 1 and PROXIMA 2A) for making these experiments possible. pRS416-prADH-GFP (pCLJ1101) was from S. Leon (Institut Jacques Monod, Paris, France), and pHCMM4 was from H. C. Medina-Munoz and M. Wickens (University of Wisconsin-Madison, Madison, USA; Addgene plasmid number 170359). This work has benefited from the crystallization (S. Plancqueel and A. Vigouroux) and Interactions of Macromolecules (M. Aumont-Nicaise) platforms of the Institute for Integrative Biology of the Cell supported by French Infrastructure for Integrated Structural Biology ANR-10-INBS-05. This study was supported by the Centre national de la recherche scientifique, France, and grant ANR-20-CE13-0007 from the Agence nationale de la recherche, France, to J.M. and C.L.J. and the Life Environment Research Center Ostrava project number CZ.10.03.01/00/22_003/0000003 via the Operational Programme Just Transition to M.E. Research in the Dacks Lab is supported by grants from the Natural Sciences and Engineering Research Council of Canada (RES0043758 and RES0046091) to J.B.D.

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Authors and Affiliations

  1. Department of Biology and Ecology, Faculty of Science, University of Ostrava, Ostrava, Czech Republic

    Romana Vargová & Marek Eliáš

  2. Université Paris-Saclay, CEA, CNRS, Institute for Integrative Biology of the Cell (I2BC), Gif-sur-Yvette, France

    Roxanne Chevreau & Julie Ménétrey

  3. Université Paris Cité, CNRS, Institut Jacques Monod, Paris, France

    Marine Alves, Camille Courbin & Catherine L. Jackson

  4. Division of Infectious Diseases, Department of Medicine, and Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada

    Kara Terry & Joel B. Dacks

  5. Synchrotron SOLEIL, l’Orme des Merisiers, Saint Aubin, France

    Pierre Legrand

  6. Institute of Parasitology, Biology Centre, Czech Academy of Sciences, České Budějovice, Czech Republic

    Joel B. Dacks

  7. Centre for Life’s Origins and Evolution, Department of Genetics, Evolution, & Environment, University College, London, UK

    Joel B. Dacks

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Contributions

R.V., R.C., M.A., C.C. and K.T. contributed to the acquisition and analysis of data; P.L. contributed to data analysis; M.E., J.M., J.B.D. and C.L.J. contributed to the conception and design of the work, data analysis and writing the manuscript.

Corresponding authors

Correspondence to Marek Eliáš, Julie Ménétrey, Joel B. Dacks or Catherine L. Jackson.

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The authors declare no competing interests.

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Nature Microbiology thanks Caner Akil, Daniel Tamarit and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data

Extended Data Fig. 1 Initial phylogenetic analysis that pointed to the existence of ArfR, a group of asgardarchaeote GTPases specifically related to eukaryotic Arf family and SRβ.

The tree was inferred using IQ-TREE with the LG + F + I + G4 model. Numbers at branches (shown when >80/95) correspond to SH-aLRT support values (right side of the slash) and ultrafast bootstrap support values (left side of the slash) calculated from 10000 replicates. As the basis for this analysis we used the sequence set utilized previously by Klinger et al.36, which included the Ras superfamily GTPase set encoded by a single asgardarchaeote MAG (the very first reported, that is ‘Lokiarchaeota archaeon GC14_75’). The dataset was expanded by addition of Ras superfamily sequences from new genome assemblies from the Asgard superphylum7,38, the korarchaeian Candidatus Korarchaeum cryptofilum, and three related sequences of an unknown provenance identified in unbinned metagenome assemblies at the early stage of the project (most likely corresponding to fragments derived from Lokiarchaeales genomes). EF-Tu sequences (GTPases not belonging to the Ras superfamily) were considered as an outgroup. For the sake of simplicity, GTPase clades observed and defined in the previous study36 were collapsed as triangles. The clade denoted ‘Ras-like’ contains eukaryotic Rab/Ras/Rho/Ran sequences, prokaryotic Rup1 sequences, and asgardarchaeote RasL as defined by Klinger et al.36. The previously undescribed group denoted ‘ArfR’ (Arf-related) consists of archaeal GTPases specifically related to eukaryotic Arf, Arl, Sar1 and SRβ proteins, whereas Rup3 denotes another previously undescribed GTPase group specific for Heimdallarchaeia (see Supplementary Fig. 7 for a more detailed analysis). Note that the existence of ArfR was missed in Klinger et al.36, because the single ArfR protein encoded by Lokiarchaeota archaeon GC14_75 (Lokiarch_19980; GenBank KKK44255.1) was removed from analysis for being considered too divergent.

Extended Data Fig. 2 Sequence diversity in the Ras superfamily analysed with CLANS.

Displayed are three different 2D projections of a 3D plot showing the results of clustering of 2994 Ras superfamily protein sequences (trimmed to the GTPase domain) based on their sequence similarity scored by all-to-all blastp comparisons. Individual sequences are represented by vertices (dots) connected by edges when their high scoring segment pairs value is ≤ 10−5. Reference sequences from different predefined subgroups are highlighted with a subgroup-specific color (according to the graphical legend provided in the middle panel). Note the clear separation from other GTPase subgroups of a cluster comprised of archaeal ArfR sequences and eukaryotic Arf family and SRβ sequences, with some of the more divergent eukaryotic sequences (for example Arl16 or SRβ) situated further away from the core of the cluster towards the periphery. Similarly, the cluster comprised of archaeal (asgardarchaeote) and eukaryotic RagGtr sequences includes some more divergent members connecting to the core from a distance. Note also the tight packing of the eukaryotic Rab/Ras/Rho/Ran sequences and the prokaryotic Rup1 group with the asgardarchaeote RasL sequences and the separate status of the newly defined Rup3 group. The analysis presented in this figure omits asgardarchaeote GTPases that remained unannotated, were annotated only based on BLAST, or were flagged as divergent (see Supplementary Table 3). An analogous clustering analysis that includes all identified asgardarchaeote Ras superfamily GTPases plus the archaeal ArvA group is displayed in Supplementary Fig. 2.

Extended Data Fig. 3 Comparison of ArfR and eukaryotic Arf family sequences.

a, Sequence alignment of HodArfR1 and GerdArfR1 with three eukaryotic Arf proteins (Homo sapiens Arf1, HsArf1, Homo sapiens Arl8A, HsArl8A and Leishmania major Arl1, LmArl1). These representatives were chosen because of structural data available (GDP-bound forms with the N-terminal AH present, in addition to the GTP-bound forms), and because they have different properties, representing the wide range of Arf family proteins in eukaryotes. The N-terminal AH is highlighted in yellow with the glycine at position 2 in black. Switch I and Switch II are highlighted in dark grey and the interswitch in light grey. The G (guanine-binding) motifs are indicated in bold and underlined. The S/T residue that (mutated to N in the dominant negative) and the Q (mutated to L in the constitutively active) mutants are indicated in red and green, respectively. Other highly conserved residues, invariant among the GTPases shown here, are indicated in black. AH, amphipathic helix; SwI, Switch I; iSw, interswitch region; SwII, Switch II. b, Helical wheel representations generated by Heliquest79 of the N-terminal AHs of the ArfR and eukaryotic Arf family sequences shown in part a). Color code for the amino acids is yellow: Val, Leu, Ile, Met, Phe, and Trp, pink: Ser and Thr, red: Asp and Glu, blue: Lys and Arg, grey: other residues. Size of letter for each amino acid in the helical wheel representation is proportional to amino acid volume; downward pointing arrow represents the helical hydrophobic moment <µH > , with length proportional to the calculated value and direction indicating that the helix is amphipathic in the perpendicular direction.

Extended Data Fig. 4 Asgardarchaeote ArfR proteins localize to the ER in yeast cells.

a, Control experiment showing that Sec13-mRFP localizes to the ER and ER exit sites. The ER protein Sec63-GFP, a component of the ER protein translocation channel, was expressed in in the yeast strain EY0987 Sec13::mRFP MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0 Sec13::mRFP-kanMX6, in which Sec13 is fused to mRFP at its C-terminus in its chromosomal locus82. Sec63-GFP (left panel), the COPII subunit Sec13-mRFP (centre panel) and the merged image (left panel) are shown. Sec13-mRFP labels both the ER and ER exit sites, which are localized as discrete puncta along the ER membrane. b, HodArfR1 and GerdArfR1 localize to the ER, marked by Sec13-mRFP. Wild type HodArfR1, HodArfR1-Q72L, wild type GerdArfR1 and GerdArfR1-Q69L were expressed as GFP fusion proteins in the yeast strain EY0987. Representative images (from three independent transformation experiments) of the indicated GFP-tagged ArfR proteins (left panels) and the same cells expressing Sec13-mRFP (centre panels) are shown. The merge image is shown in the right panels. c, HodArfR1 and GerdArfR1 localize to the ER, marked by Sec61-mCherry, the ER protein translocation channel. Wild type HodArfR1 and wild type GerdArfR1 were expressed as GFP fusion proteins in the yeast strain BY4742 MATα his3Δ1 leu2Δ0 lys2Δ0 ura3Δ0, which was co-transformed with pHCMM4 Sec61-mCherry. Representative images (from at least three independent transformation experiments) of the indicated GFP-tagged ArfR proteins (left panels) and the same cells expressing Sec61-mCherry (centre panels) are shown. The merge image is shown in the right panels.

Extended Data Fig. 5 GDP/GTP structural cycle of GerdArfR1.

a, (left) Crystal structure of GerdArfR1-Δ12-Q69L bound to Mg.GTP. The protein is shown with the interswitch in blue and Switch I/Switch II in light blue. Below, the nucleotide binding site of the Mg.GTP is enlarged. The N-terminal AH which is absent from the fragment crystallized, is shown schematically. (right) Crystal structure of GerdArfR1-FL-WT bound to Mg.GDP. The protein is shown with the N-terminal helix and the interswitch in pink and Switch I/Switch II in light pink. Above, the N-terminal helix is magnified and shown in two orientations rotated by 90°. Hydrophobic residues Leu3, Phe5, Leu6, Leu9 and Phe10 from the N-terminal AH of GerdArfR1 (indicated in yellow) are buried in the hydrophobic pocket formed by the tip of the interswitch and the C-terminal helix. Overall, proteins are shown with a cartoon representation, magnesium ions with a green sphere and nucleotides with a stick representation. b, Superposition of the GTP-bound and GDP-bound forms of HodArfR1. Colors are as described in part a.

Extended Data Fig. 6 Crystal dimer of GerdArfR1-GDP.

a, Overall organization of the crystal dimer of GerdArfR1-GDP. The N-terminal AH and switch regions of molecule A are in pink and those of molecule B are in orange. b-c, Detailed views of the two interfaces of the dimer involving Switch II of one molecule and the β2-strand of the interswitch from the other molecule. d, The hydrogen network of the interswitch (β2-β3 strands) shows an unzipping compared to other Arf-GDP structures. e, The atypical coordination sphere of the magnesium ion. The classical position of the magnesium ion is indicated by a transparent green sphere. f, Sec-RALS experiment showing that in solution GerdArfR1-GDP is monomeric.

Extended Data Fig. 7 Structural comparisons of the GDP-bound forms of asgardarchaeote and eukaryotic Arf proteins.

HodArfR1-GDP, GerdArfR1-GDP, HsArf1-GDP, HsArl8-GDP and LmArl1-GDP structures are compared to each other in pairs. A face orientation shows the N-terminal and the interswitch regions in colors and Switch I/Switch II regions in grey (upper part of each panel) and a 90° orientation shows the N-terminal helix (lower part of each panel). Superposition was performed on residues of the P-loop, β1-β3 strands and the C-terminal helix. The same orientation is shown for all superpositions. Hod, Hodarchaeales; Hs, Homo sapiens; Lm, Leishmania major. Pairwise comparisons of full Arf family protein structures show that RMSD (using Cα atoms) ranges from 1.5 Å to 4.0 Å and decreases from 0.9 Å to 1.9 Å when N-terminal AH and Switch I/Switch II sequences are removed (noted hereafter as RMSD 1.5-4.0/0.9-1.9 Å). HsArf1 and LmArl1 are structurally very close (RMSD 1.5/0.9 Å), while the difference between human Arf1 and Arl8A is greater (RMSD 2.9/1.3 Å), but similar to that observed between archaeal HodArfR1 and the three eukaryotic Arf family protein structures (2.6-2.8/1.2-1.3 Å) or between archaeal GerdArfR1 and HsArf1/LmArl1 structures (RMSD 2.8-3.0/1.4-1.7 Å). One difference comes from pairwise comparison between archaeal GerdArfR1 and HsArl8, which exhibit larger RMSDs of 3.6/1.9 Å, similar to those measured between HodArfR1 and GerdArfR1 (RMSD 4.0/1.9 Å).

Supplementary information

Supplementary Information

Supplementary Figs. 1–9, Notes 1–6 and refs. 1–13.

Reporting Summary

Supplementary Tables

Supplementary Tables 1–10.

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Vargová, R., Chevreau, R., Alves, M. et al. The Asgard archaeal origins of Arf family GTPases involved in eukaryotic organelle dynamics. Nat Microbiol 10, 495–508 (2025). https://doi.org/10.1038/s41564-024-01904-6

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