Article | Published:

A plasmid from an Antarctic haloarchaeon uses specialized membrane vesicles to disseminate and infect plasmid-free cells


The major difference between viruses and plasmids is the mechanism of transferring their genomic information between host cells. Here, we describe the archaeal plasmid pR1SE from an Antarctic species of haloarchaea that transfers via a mechanism similar to a virus. pR1SE encodes proteins that are found in regularly shaped membrane vesicles, and the vesicles enclose the plasmid DNA. The released vesicles are capable of infecting a plasmid-free strain, which then gains the ability to produce plasmid-containing vesicles. pR1SE can integrate and replicate as part of the host genome, resolve out with fragments of host DNA incorporated or portions of the plasmid left behind, form vesicles and transfer to new hosts. The pR1SE mechanism of transfer of DNA could represent the predecessor of a strategy used by viruses to pass on their genomic DNA and fulfil roles in gene exchange, supporting a strong evolutionary connection between plasmids and viruses.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


  1. 1.

    Huber, H. et al. A new phylum of Archaea represented by a nanosized hyperthermophilic symbiont. Nature 417, 63–67 (2002).

  2. 2.

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

  3. 3.

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

  4. 4.

    Wang, H., Peng, N., Shah, A. S., Huang, L. & She, Q. Archaeal extrachromosomal genetic elements. Microbiol. Mol. Biol. Rev. 79, 117–152 (2015).

  5. 5.

    Forterre, P. & Krupovic, M. in Viruses: Essential Agents of Life (Ed. Witzany, G.) 43–60 (Springer, Dordrecht, 2012).

  6. 6.

    Holmes, E. C. What does virus evolution tell us about virus origins? J. Virol. 85, 5247–5251 (2011).

  7. 7.

    Gorlas, A., Krupovic, M., Forterre, P. & Geslin, G. Living side by side with a virus: characterisation of two novel plasmids from Thermococcus prieurii, a host for the spindle-shaped virus TPV1. Appl. Environ. Microbiol. 79, 3822–3828 (2013).

  8. 8.

    Cavicchioli, R. Microbial ecology of Antarctic aquatic systems. Nat. Rev. Microbiol. 13, 691–706 (2015).

  9. 9.

    Forterre, P., Krupovic, M., Raymann, K. & Soler, N. Plasmids from Euryarchaeota. Microbiol. Spectr2, PLAS-0027-2014 (2014).

  10. 10.

    Arnold, H. P. et al. The genetic element pSSVx of the extremely thermophilic crenarchaeon Sulfolobus is a hybrid between a plasmid and a virus. Mol. Microbiol. 34, 217–226 (1999).

  11. 11.

    Ye, X., Ou, J., Ni, L., Shi, W. & Shen, P. Characterization of a novel plasmid from extremely halophilic Archaea: nucleotide sequence and function analysis. FEMS. Microbiol. Lett. 221, 53–57 (2003).

  12. 12.

    Zhang, Z. et al. Temperate membrane-containing halophilic archaeal virus SNJ1 has a circular dsDNA genome identical to that of plasmid pHH205. Virology 434, 233–241 (2012).

  13. 13.

    Gaudin, M. et al. Extracellular membrane vesicles harbouring viral genomes. Environ. Microbiol. 16, 1167–1175 (2014).

  14. 14.

    Krupovic, M. & Bamford, D. H. Order to the viral universe. J. Virol. 84, 12476–12479 (2010).

  15. 15.

    Naor, A., Lapierre, P., Mevarech, M., Papke, R. T. & Gophna, U. Low species barriers in halophilic archaea and the formation of recombinant hybrids. Curr. Biol. 22, 1444–1448 (2012).

  16. 16.

    Koonin, E. V. & Dolja, V. V. Virus world as an evolutionary network of viruses and capsidless selfish elements. Microbiol. Mol. Biol. Rev. 78, 278–303 (2014).

  17. 17.

    Krupovic, M. Recombination between RNA viruses and plasmids might have played a central role in the origin and evolution of small DNA viruses. Bioessays 34, 867–870 (2012).

  18. 18.

    Deatherage, B. L. & Cookson, B. T. Membrane vesicle release in bacteria, eukaryotes, and archaea: a conserved yet underappreciated aspect of microbial life. Infect. Immun. 80, 1948–1957 (2012).

  19. 19.

    Biller, S. J. et al. Bacterial vesicles in marine ecosystems. Science 343, 183–186 (2014).

  20. 20.

    Soler, N., Krupovic, M., Marguet, E. & Forterre, P. Membrane vesicles in natural environments: a major challenge in viral ecology. ISME J. 9, 793–796 (2015).

  21. 21.

    DeMaere, M. Z. et al. High level of inter-genera gene exchange shapes the evolution of haloarchaea in an isolated Antarctic lake. Proc. Natl. Acad. Sci. USA 110, 16939–16944 (2013).

  22. 22.

    Tschitschko, B. et al. Antarctic archaea–virus interactions: metaproteome-led analysis of invasion, evasion and adaptation. ISME J. 9, 2094–2107 (2015).

  23. 23.

    Franzmann, P. D. et al. Halobacterium lacusprofundi sp. nov., a halophilic bacterium isolated from Deep Lake, Antarctica. Syst. Appl. Microbiol. 11, 20–27 (1988).

  24. 24.

    Ng, W. L. & DasSarma, S. Minimal replication origin of the 200-kilobase Halobacterium plasmid pNRC100. J. Bacteriol. 175, 4584–4596 (1993).

  25. 25.

    Gotfredsen, M. & Gerdes, K. The Escherichia coli relBE genes belong to a new toxin–antitoxin gene family. Mol. Microbiol 29, 1065–1076 (1998).

  26. 26.

    Xu, C. & Min, J. Structure and function of WD40 domain proteins. Protein Cell 2, 202–214 (2011).

  27. 27.

    Faini, M., Beck, R., Wieland, F. T. & Briggs, J. A. Vesicle coats: structure, function, and general principles of assembly. Trends Cell. Biol. 23, 279–288 (2013).

  28. 28.

    Gürkan, C., Stagg, S. M., LaPointe, P. & Balch, W. E. The COPII cage: unifying principles of vesicle coat assembly. Nat. Rev. Mol. Cell Biol. 7, 727–738 (2006).

  29. 29.

    Ter Haar, E., Musacchio, A., Harrison, S. C. & Kirchhausen, T. Atomic structure of clathrin: a β propeller terminal domain joins an α zigzag linker. Cell 95, 563–573 (1998).

  30. 30.

    Santarella-Mellwig, R. et al. The compartmentalized bacteria of the Planctomycetes–Verrucomicrobia–Chlamydiae superphylum have membrane coat-like proteins. PLoS Biol. 8, e1000281 (2010).

  31. 31.

    Alber, F. et al. The molecular architecture of the nuclear pore complex. Nature 450, 695–701 (2007).

  32. 32.

    Quemin, E. R. & Quax, T. E. Archaeal viruses at the cell envelope: entry and egress. Front. Microbiol 6, 552 (2015).

  33. 33.

    Shaik, M. M. et al. A structural snapshot of type II pilus formation in Streptococcus pneumoniae. J. Biol. Chem. 290, 22581–22592 (2015).

  34. 34.

    Duggin, I. et al. CetZ tubulin-like proteins control archaeal cell shape. Nature 519, 362–365 (2015).

  35. 35.

    Knowles, T. J., Scott-Tucker, A., Overduin, M. & Henderson, I. R. Membrane protein architects: the role of the BAM complex in outer membrane protein assembly. Nat. Rev. Microbiol 7, 206–214 (2009).

  36. 36.

    Lee, E. Y. et al. Global proteomic profiling of native outer membrane vesicles derived from Escherichia coli. Proteomics 7, 3143–3153 (2007).

  37. 37.

    Barlowe, C., d’Enfert, C. & Schekman, R. Purification and characterization of SAR1p, a small GTP-binding protein required for transport vesicle formation from the endoplasmic reticulum. J. Biol. Chem. 268, 873–879 (1993).

  38. 38.

    Serafini, T. et al. ADP ribosylation factor is a subunit of the coat of Golgi-derived COP-coated vesicles: a novel role for a GTP-binding protein. Cell 67, 239–253 (1991).

  39. 39.

    Goessweiner-Mohr, N., Arends, K., Keller, W. & Grohmann, E. Conjugative type IV secretion systems in Gram-positive bacteria. Plasmid 70, 289–302 (2013).

  40. 40.

    Krupovic, M., Ravantti, J. J. & Bamford, D. H. Geminiviruses: a tale of a plasmid becoming a virus. BMC Evol. Biol. 9, 112 (2009).

  41. 41.

    Jalasvuori, M., Mattila, S. & Hoikkala, V. Chasing the origin of viruses: capsid-forming genes as a life-saving preadaptation within a community of early replicators. PLoS ONE 8, e0126094 (2015).

  42. 42.

    Cantin, R., Methot, S. & Tremblay, M. J. Plunder and stowaways: incorporation of cellular proteins by enveloped viruses. J. Virol. 79, 6577–6587 (2005).

  43. 43.

    Shaw, M. L., Stone, K. L., Colangelo, C. M., Gulcicek, E. E. & Palese, P. Cellular proteins in influenza virus particles. PLoS Pathog. 4, e1000085 (2008).

  44. 44.

    Sun, E., He, J. & Zhuang, X. Dissecting the role of COPI complexes in influenza virus infection. J. Virol. 87, 2673–2685 (2013).

  45. 45.

    Pietila, M. K. et al. Virion architecture unifies globally distributed pleolipoviruses infecting halophilic archaea. J. Virol. 86, 5067–5079 (2012).

  46. 46.

    Dyall-Smith, M. The Halohandbook: Protocols for Halobacterial Genetics (Haloarchaea and Haloviruses, 2008);

  47. 47.

    Williams, T. J. et al. Microbial ecology of an Antarctic hypersaline lake: genomic assessment of ecophysiology amongst dominant haloarchaea. ISME J. 8, 1645–1658 (2014).

  48. 48.

    Liao, Y. et al. Developing a genetic manipulation system for the Antarctic archaeon, Halorubrum lacusprofundi: investigating acetamidase gene function. Sci. Rep. 6, 34639 (2016).

  49. 49.

    Campbell, P. J. Primary productivity of a hypersaline Antarctic lake. Mar. Freshwat. Res. 29, 717–724 (1978).

  50. 50.

    Ferris, J. M. & Burton, H. R. The annual cycle of heat content and mechanical stability of hypersaline Deep Lake, Vestfold Hills, Antarctica. Hydrobiologia 165, 115–128 (1988).

  51. 51.

    Nurk, S. et al. in Research in Computational Molecular Biology (eds Deng, M., Jiang, R., Sun, F. & Zhang, X.) 158–170 (Springer, Berlin, Heidelberg, 2013).

  52. 52.

    Nurk, S., Meleshko, D., Korobeynikov, A. & Pevzner, P. metaSPAdes: a new versatile de novo metagenomics assembler. Preprint at (2016).

  53. 53.

    Overbeek, R. et al. The SEED and the Rapid Annotation of microbial genomes using Subsystems Technology (RAST). Nucleic Acids Res. 42, D206–D214 (2014).

  54. 54.

    Altschul, S. F., Gish, W., Miller, W., Myers, E. W. & Lipman, D. J. Basic local alignment search tool. J. Mol. Biol. 215, 403–410 (1990).

  55. 55.

    Mitchell, A. et al. The InterPro protein families database: the classification resource after 15 years. Nucleic Acids Res. 43, 213–221 (2015).

  56. 56.

    Söding, J. Protein homology detection by HMM-HMM comparison. Bioinformatics 21, 951–960 (2005).

  57. 57.

    Yang, J. et al. The I-TASSER Suite: protein structure and function prediction. Nat. Methods 12, 7–8 (2015).

  58. 58.

    Carver, T., Thomson, N., Bleasby, A., Berriman, M. & Parkhill, J. DNAPlotter: 503 circular and linear interactive genome visualization. Bioinformatics 25, 119–120 (2009).

  59. 59.

    Carver, T., Harris, S. R., Berriman, M., Parkhill, J. & McQuillan, J. A. Artemis: an integrated platform for visualization and analysis of high-throughput sequence-based experimental data. Bioinformatics 28, 464–469 (2012).

  60. 60.

    Michalski, A. et al. Mass spectrometry-based proteomics using Q Exactive, a high-performance benchtop quadrupole Orbitrap mass spectrometer. Mol. Cell. Proteomics 10, M111.011015 (2011).

  61. 61.

    Ishihama, Y. et al. Exponentially modified protein abundance index (emPAI) for estimation of absolute protein amount in proteomics by the number of sequenced peptides per protein. Mol. Cell. Proteomics 4, 1265–1272 (2005).

  62. 62.

    Arike, L. & Peil, L. Spectral counting label-free proteomics. Methods Mol. Biol. 1156, 213–222 (2014).

  63. 63.

    Ellen, A. F. et al. Proteomic analysis of secreted membrane vesicles of archaeal Sulfolobus species reveals the presence of endosome sorting complex components. Extremophiles 13, 67–79 (2009).

  64. 64.

    Vizcaíno, J. A. et al. ProteomeXchange provides globally co-ordinated proteomics data submission and dissemination. Nat. Biotechnol. 30, 223–226 (2014).

  65. 65.

    Wu, Z., Liu, H., Liu, J., Liu, X. & Xiang, H. Diversity and evolution of multiple orc/cdc6-adjacent replication origins in haloarchaea. BMC Genomics 13, 478 (2012).

Download references


This work was supported by the Australian Research Council (DP150100244) and the Australian Antarctic Science program (project 4031). S.E. was supported by the EMBO Long-Term Fellowship ALTF 188–2014, which is co-funded by the European Commission (EMBOCOFUND2012, GA-2012-600394) and supported by Marie Curie Actions. Mass spectrometry results were obtained at the Bioanalytical Mass Spectrometry Facility (BMSF) and electron microscopy at the Electron Microscope Unit, both within the Analytical Centre of the University of New South Wales. Subsidized access to the BMSF is acknowledged. The authors thank the PRIDE team and ProteomeXchange for efficiently processing and hosting the mass spectrometry data. The authors thank A. Hancock for providing the image of Rauer 1 Lake, the Landsat Image Mosaic of Antarctica (LIMA) project for making satellite images available, S. Payne and A. Hancock for collecting Antarctic water samples, M. Allen for providing uncharacterized strains of Hrr. lacusprofundi, R. Kuchel for assistance with electron microscopy, D. Baker and I. Anishchanka for attempting structural predictions and T. Williams for comments about the manuscript.

Author information

S.E. and R.C. conceived and led the study and performed the primary writing of the manuscript. S.E. performed all experimental work related to VLPs and host strains, including discovering the existence of PVs. B.T. assembled DNA sequence data and analysed metagenome data. L.Z. and M.J.R. performed the mass spectrometry. All authors participated in the analysis and interpretation of the data and contributed to the writing of the manuscript.

Competing interests

The authors declare no competing financial interests and no conflict of interest.

Correspondence to Ricardo Cavicchioli.

Electronic supplementary material

Supplementary Information

Supplementary Notes, Supplementary References, Supplementary Figures and Supplementary Tables.

Supplementary Data File 1

pR1SEDL18 contig with annotation

Supplementary Data File 2

All proteins detected in vesicle and membrane preparations.

Supplementary Date File 3

All proteins detected in PVs purified by CsCl gradient centrifugation.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Further reading

Fig. 1: Transmission electron micrographs of Hrr. lacusprofundi R1S1 VLPs.
Fig. 2: Plasmid map of pR1SE.
Fig. 3: Functional predictions of pR1SE proteins from region 2 that were detected in PVs.
Fig. 4: pR1SE derivatives in Hrr. lacusprofundi hosts.
Fig. 5: Overview of the PV ‘life cycle’: PV formation and infection, and pR1SE dissemination, integration and resolution.