Introduction

Cenancestor stands for the last universal cellular ancestor (LUCA) of all living cellular organisms, where the Greek prefix cen- means recent (as in Cenozoic Era) and common or shared (as in cœnocyte). The cenancestor here is neither the initial Darwinian ancestor in the RNA world [1] which had not yet gained a cellular form, nor the protocell that marked the beginning of cellular life [2] which might not contain inheritable genetic material. The cenancestor could have lived with many other cellular organisms, except that all other cellular organisms failed to leave any descendants in today’s biosphere.

There is overwhelming evidence that all living cellular forms of life, represented by Archaea, Eubacteria and Eukarya, are genetically related. First, the genetic code is almost universally conserved. Second, different genes from representatives of these three kingdoms of life often yield similar phylogenetic relationships. Third, all cellular forms of life share substantial similarity in the three most fundamental biological processes (genome replication, transcription and translation). Fourth, the common ancestry has been substantiated by numerous transitional fossils. Fifth, evolutionary convergence observed either at the phenotypic or at the DNA level, provides no sufficient explanation for the phenotypic or genetic similarities among organisms. Finally, if the cellular structure originated only once, then the existence of a cellular cenancestor is a logical necessity given the cell theory developed by Theodor Schwann, Matthias Jacob Schneider, and Rudolf Virchow in the late 17^th century which states that news cells are created by old cells dividing into two. However, the empirical search for the cenancestor started only in 1977 and brought about fundamental changes in biology, especially in evolutionary biology.

The prediction and discovery of an ancient lineage

In early 1977, Woese and Fox [3] noted many differences in translation machinery between Bacteria and Eukarya. For example, the 5S RNA in Bacteria has no counterpart in Eukarya, and the 5.8S RNA in Eukarya has no counterpart in Bacteria. High interchangeability of components of the translation machinery is observed within Bacteria or within Eukarya, but not between the two. Bacterial lineages have evolved for billions of years, yet their translation machineries are highly conserved. If the common ancestor of bacterial species is an ensemble of extant lineages, then the translation machinery of this bacterial common ancestor is very unlikely to evolve into a eukaryotic one. Their hypothesis is that Eukarya must have evolved from a primitive lineage quite different from bacteria. There must be an ancient "progenote" lineage that predates the common ancestor of the extant bacterial lineages and gave rise to eukaryotes.

In late 1977, Woese and Fox [4] presented their evidence for the three primary domains of life, with Archaebacteria as a new domain potentially representing the extant lineages of the progenote. They generated a similarity matrix of the three domains, but did not build a tree or speculate on which two domains were more closely related – the three domains appeared equidistant when a distance-based tree was reconstructed from their matrix [5]. The cenancestor was implicitly assumed to have trifurcated to give rise to the three primary domains of life. This implicit trifurcation might have contributed to the ill-fated paleocyte hypothesis [6] and the original Neomera hypothesis [7-9], both hypotheses assuming that Eukarya and Archaebacteria are monophyletic and sister to each other.

Identification of the root of the three domains of life

Three approaches were immediately used to detect phylogenetic relationships among the three domains of life in an effort to identify the root, which is an approximation of the cenancestor. The first is by similarities in organellar structures and gene sharing. Ribosome structure provided the first indication that Archaebacteria and Eukarya are more similar to each other than they are to Eubacteria [6,10]. However, one can easily find counter examples. For example, gluthamine synthetase (GS) has three types, GSI, GSII and GSIII. GSI homologues have been found only in Eubacteria and Archaea, and GSII genes only in eukaryotes and a few bacterial lineages [11]. If gene-sharing is a strong indicator of phylogenetic affinity, then GSI would favour the grouping of Eubacteria and Archaea, but GSII would favour the grouping of Eukarya and Eubacteria.

The second approach to identify the root is by molecular phylogenetics enabled by two advances. The first is the recognition of a set of genes as reliable genetic markers. These genes are involved DNA replication [12,13], transcription [14-17] and translation [18-20]. As they participate in the transmission of genetic information, they were subsequently named informational genes [21,22]. In particular, informational genes are highly conservative (suitable for resolving deep phylogenies) and rarely involved in horizontal gene transfer [21-23], although aminoacyl-tRNA synthetases are known to be horizontally transferred [24]. The second, and more important, advance is the innovative "rooting by paralogue" strategy [25,26] depicted in Fig. 1, where one paralogue can be used to root the tree of the other. The approach favoured the tree in Fig. 1C. Furthermore, it placed Eukarya as a clade within the Archaebacteria domain, with Eukarya as a sister group of Crenarchaeota [27-29]. Subsequent refinements with more sequences and less uncertainty in orthology identification clustered Eukarya with the TACK superphylum [30] or Lokiarchaeota [31,32] within Crenarchaeota. Overwhelming evidence suggests that Archaebacteria was "archae" only to eukaryotes, but not to Eubacteria. Consequently, Archaebacteria was renamed to Archaea.

Fig. 1. Testing three alternative hypotheses with the "rooting by paralogue" approach. A gene duplication event occurred before divergence among the three domains of life, leading to two paralogues (G1 and G2) in each domain. (A) Monophyly of prokaryotes. (B) Eukarya more closely related to Eubacteria than to Archaea. (C) Eukarya and Archaea are sister lineages.

The third approach to identify the root is by character polarization [7-9,33] which is often used in evolutionary inference. For example, the nuclear organization of Eukarya is reasonably assumed to be a derived character relative to the lack of such an organization in prokaryotes, and multicellularity is a derived character relative to unicellularity, although polarization of characters can sometimes change direction. Three key features are considered as derived characters in the context of the three domains of life: 1) chromosome packaging with histones, 2) methionine as initiator amino acid in translation (in contrast to formylmethionine in Eubacteria), and 3) multiple RNA polymerases (in contrast to Eubacteria with only one). Archaea and Eukarya generally share all these derived characters, which led to the proposal of the Neomura hypothesis [7-9] in which Neomura includes Archaea and Eukarya as mutual sisters. There are also some other characters that are predominantly but not uniquely neomuran, such as the use of cholesterols and proteasomes [7]. However, these "derived characters" are not necessarily conserved and could be lost and gained multiple times. For example, within Crenarchaeota alone, there are lineages with or without histones [34]. This original Neomura hypothesis with Archaea and Eukarya as sisters has now been discarded as it is not supported even by its original advocates [35]. There are other derived characteristics. For example, RNA ribozymes occur in the RNA world [36] before the appearance protein enzymes [36,37]. Similarly, ribosome RNAs should form the primitive translation machinery in the RNA world before participation of ribosome proteins and translation factors [38]. However, the cenancester is assumed to appear long after the RNA world.

With increasing number of genes used in resolving the phylogeny of the three domains, it has become clear that the relationship is volatile and depends on what genes one chooses to characterize the relationship. For example, although most genes involved in translation tend to group Archaea and Eukarya together, glutamine synthetase sequences would unambiguously cluster the archaea and Eubacteria together to the exclusion of Eukarya. Such cases frequently force researchers to invoke horizontal gene transfer as a post hoc explanation [11,39]. All these observations of conflicting phylogenetic signals suggest a mosaic nature of the ancestral eukaryotic genome, i.e., it has incorporated both Eubacteria and Archaea genes.

Extensive search for the cenancestor has reduced the three domains of life to two domains [40,41], with Eukarya nested within Archaea and the cenancestor is rooted between Archaea and Eubacteria. Extensive horizontal gene transfer has contributed to the evolution of Eukarya leading to the mosaic nature of the eukaryotic genome. In this context, the domain including Archaea and Eukarya should be renamed to Eukaryoida, with Eukarya renamed to Eukaryae as a kingdom within the Eukaryoida domain.

Endosymbiotic hypothesis of organellar origin

The conceptual formulation [42-44] and molecular substantiation of the endosymbiotic hypothesis for the origin of mitochondria [45-48] and chloroplasts [49,50] shed lights on the origin and evolution of the first eukaryote. The eukaryotic nucleus itself could be the evolutionary consequence of endosymbiosis. If a prokaryotic cell (a progenote) engulfed an aerobic bacterial cell [3] or vice versa [6], then the DNA of the engulfing host could evolve towards a nucleus and the DNA of the engulfed guest would evolve towards mitochondria or chloroplasts. Both the host genome and the guest genome would contribute to the final nuclear genome, leading to the mosaic nature of the eukaryotic genome that continue to frustrate phylogeneticists with its different genes generating different trees. Gene transfer from mitochondria to nucleus was long suspected [44,51] and now comprehensively verified [52,53]. Such transfer are particularly common in plants [54-57], and has occurred early, close to the root of plants [58].

Eukaryotic origin and evolution: the engulfing host and the engulfed guest

The mitochondrial progenitor is now generally accepted to be a Rickettsia-like bacterium [59,60] internalized most likely into a prokaryotic cell (Gray, 2012; Lane and Martin, 2010). Which lineage serves as the engulfing host remains controversial. However, recent studies [41,61] provides compelling evidence for a close relationship between Eukarya and Asgard archaea. It is reasonable to hypothesize that an Asgard archaeal cell engulfed an aerobic Rickettsia-like bacterium, leading to the reticulate origin of eukaryotes. The genome of the engulfing host serves as the beginning of the eukaryotic nuclear genome, and the engulfed bacterium evolved into mitochondria. Extensive horizontal gene transfer from the mitochondrial genome to the nuclear genome gave rise to the mosaic nature of the eukaryotic genome.

One difficulty with the hypothesis of eukaryotic origin above is that prokaryotes with a rigid cell wall could not engulf one another, so nature would need a wall-less archaea to serve as the engulfing host. Thermoplasma and Ferroplasma are wall-less Euryarchaeota [62]. However, they are not closely related to Eukarya as Asgard archaea.

Viruses and plasmids: the origin of endosymbiotic genomes without the need of cells engulfing each other

I have so far ignored viruses, but two lines of evidence suggest the possibility that viruses coexisted with the cenancestor. First, viruses have at least four realms that do not share a common ancestor [63]. Second, and giant viruses harbour protein domains not known in cellular forms of life [64]. Viruses could have mediated horizontal gene transfer among the early lineages of life and bring heterogeneous genomes together, which would abolish the need of primitive cells engulfing each other to create endosymbiotic genomes. Plasmids could make similar contributions in bringing different genomes into the same cell.

Given the generally accepted reticulate speciation that gave rise to eukaryotes, the extensive horizontal gene transfer among ancient lineages, and the coexistence of viruses that would facilitate different genomes coming together for genetic exchange, one should not view the cenancestor as a single cell or a single genome. Instead, the cenancestor is better perceived as a pool of promiscuous cellular forms of life with relatively free exchange of genetic materials leading to frequent reticulate speciation events. Out of this pool of frolicking genomes arose many domains of life of which only Eubacteria and Eukaryoida (Archaea + Eukarya) have extant lineages today.

References

1. Yarus, M., Getting Past the RNA World: The Initial Darwinian Ancestor. In RNA World: From life's origin to diversity in gene regulation, Atkins, J.F.; Gesteland, R.F.; Cech, T.R., Eds. Cold Spring Harbor Laboratory Press: Cold Spring Harbor Laboratory, 2011; pp 43-50.
2. Schrum, J.P.; Zhu, T.F.; Szostak, J.W., The Origins of Cellular Life. In RNA World: From life's origin to diversity in gene regulation, Atkins, J.F.; Gesteland, R.F.; Cech, T.R., Eds. Cold Spring Harbor Laboratory Press: Cold Spring Harbor Laboratory, 2011; pp 51-62.
3. Woese, C.R.; Fox, G.E., The concept of cellular evolution. J Mol Evol 1977, 10, 1-6.
4. Woese, C.R.; Fox, G.E., Phylogenetic structure of the prokaryotic domain: the primary kingdoms. Proc. Natl. Acad. Sci. U S A 1977, 74, 5088-5090.
5. Xia, X., A Mathematical Primer of Molecular Phylogenetics. CRC Press: New York, 2020; p 380.
6. Lake, J.A.; Henderson, E.; Clark, M.W.; Matheson, A.T., Mapping evolution with ribosome structure: intralineage constancy and interlineage variation. Proc. Natl. Acad. Sci. U S A 1982, 79, 5948-5952.
7. Cavalier-Smith, T., The phagotrophic origin of eukaryotes and phylogenetic classification of Protozoa. Int J Syst Evol Microbiol 2002, 52, 297-354.
8. Cavalier-Smith, T., Rooting the tree of life by transition analyses. Biol Direct 2006, 1, 19.
9. Cavalier-Smith, T., The neomuran revolution and phagotrophic origin of eukaryotes and cilia in the light of intracellular coevolution and a revised tree of life. Cold Spring Harb Perspect Biol 2014, 6, a016006.
10. Lake, J.A.; E. Henderson; Oakes, M.; Clark, a.M.W., Eocytes: A new ribosome structure indicates a kingdom with a close relationship to eukaryotes. Proc. Natl. Acad. Sci. USA. 1984, 81, 3786-3790.
11. Brown, J.R.; Masuchi, Y.; Robb, F.T.; Doolittle, W.F., Evolutionary relationships of bacterial and archaeal glutamine synthetase genes. J Mol Evol 1994, 38, 566-576.
12. Cann, I.K.; Ishino, Y., Archaeal DNA replication: identifying the pieces to solve a puzzle. Genetics 1999, 152, 1249-1267.
13. Raymann, K.; Forterre, P.; Brochier-Armanet, C.; Gribaldo, S., Global phylogenomic analysis disentangles the complex evolutionary history of DNA replication in archaea. Genome Biol Evol 2014, 6, 192-212.
14. Langer, D.; Hain, J.; Thuriaux, P.; Zillig, W., Transcription in archaea: similarity to that in eucarya. Proc. Natl. Acad. Sci. U S A 1995, 92, 5768-5772.
15. Marsh, T.L.; Reich, C.I.; Whitelock, R.B.; Olsen, G.J., Transcription factor IID in the Archaea: sequences in the Thermococcus celer genome would encode a product closely related to the TATA-binding protein of eukaryotes. Proc. Natl. Acad. Sci. USA. 1994, 91, 4180-4184.
16. Qureshi, S.A.; Baumann, P.; Rowlands, T.; Khoo, B.; Jackson, S.P., Cloning and functional analysis of the TATA binding protein from Sulfolobus shibatae. Nucleic Acids Res 1995, 23, 1775-1781.
17. Wettach, J.; Gohl, H.P.; Tschochner, H.; Thomm, M., Functional interaction of yeast and human TATA-binding proteins with an archaeal RNA polymerase and promoter. Proc. Natl. Acad. Sci. U S A 1995, 92, 472-476.
18. Lechner, K.; Heller, G.; Böck, A., Gene for the diphtheria toxin-susceptible elongation factor 2 from Methanococcus vannielii. Nucleic Acids Res 1988, 16, 7817-7826.
19. Berghöfer, B.; Kröckel, L.; Körtner, C.; Truss, M.; Schallenberg, J.; Klein, A., Relatedness of archaebacterial RNA polymerase core subunits to their eubacterial and eukaryotic equivalents. Nucleic Acids Res 1988, 16, 8113-8128.
20. Creti, R.; Ceccarelli, E.; Bocchetta, M.; Sanangelantoni, A.M.; Tiboni, O.; Palm, P.; Cammarano, P., Evolution of translational elongation factor (EF) sequences: reliability of global phylogenies inferred from EF-1 alpha(Tu) and EF-2(G) proteins. Proc. Natl. Acad. Sci. U S A 1994, 91, 3255-3259.
21. Jain, R.; Rivera, M.C.; Lake, J.A., Horizontal gene transfer among genomes: the complexity hypothesis. Proc. Natl. Acad. Sci. U S A 1999, 96, 3801-3806.
22. Jain, R.; Rivera, M.C.; Moore, J.E.; Lake, J.A., Horizontal gene transfer accelerates genome innovation and evolution. Mol. Biol. Evol. 2003, 20, 1598-1602.
23. Rivera, M.C.; Jain, R.; Moore, J.E.; Lake, J.A., Genomic evidence for two functionally distinct gene classes. Proc. Natl. Acad. Sci. USA. 1998, 95, 6239-6244.
24. Dohm, J.C.; Vingron, M.; Staub, E., Horizontal gene transfer in aminoacyl-tRNA synthetases including leucine-specific subtypes. J Mol Evol 2006, 63, 437-447.
25. Iwabe, N.; Kuma, K.; Hasegawa, M.; Osawa, S.; Miyata, T., Evolutionary relationship of archaebacteria, eubacteria, and eukaryotes inferred from phylogenetic trees of duplicated genes. Proc. Natl. Acad. Sci. USA. 1989, 86, 9355-9359.
26. Gogarten, J.P.; Kibak, H.; Dittrich, P.; Taiz, L.; Bowman, E.J.; Bowman, B.J.; Manolson, M.F.; Poole, R.J.; Date, T.; Oshima, T., et al., Evolution of the vacuolar H+-ATPase: implications for the origin of eukaryotes. Proc. Natl. Acad. Sci. U S A 1989, 86, 6661-6665.
27. Baldauf, S.L.; Palmer, J.D.; Doolittle, W.F., The root of the universal tree and the origin of eukaryotes based on elongation factor phylogeny. Proc. Natl. Acad. Sci. U S A 1996, 93, 7749-7754.
28. Brown, J.R.; Robb, F.T.; Weiss, R.; Doolittle, W.F., Evidence for the early divergence of tryptophanyl- and tyrosyl- tRNA synthetases. J. Mol. Evol. 1997, 45, 9-16.
29. Brown, J.R.; Doolittle, W.F., Root of the universal tree of life based on ancient aminoacyl-tRNA synthetase gene duplications. Proc. Natl. Acad. Sci. U S A 1995, 92, 2441-2445.
30. Raymann, K.; Brochier-Armanet, C.; Gribaldo, S., The two-domain tree of life is linked to a new root for the Archaea. Proc. Natl. Acad. Sci. U S A 2015, 112, 6670-6675.
31. Spang, A.; Saw, J.H.; Jorgensen, S.L.; Zaremba-Niedzwiedzka, K.; Martijn, J.; Lind, A.E.; van Eijk, R.; Schleper, C.; Guy, L.; Ettema, T.J.G., Complex archaea that bridge the gap between prokaryotes and eukaryotes. Nature 2015, 521, 173-179.
32. López-García, P.; Moreira, D., Open Questions on the Origin of Eukaryotes. Trends Ecol Evol 2015, 30, 697-708.
33. Cavalier-Smith, T., Deep phylogeny, ancestral groups and the four ages of life. Philos Trans R Soc Lond B Biol Sci 2010, 365, 111-132.
34. Cubonová, L.; Sandman, K.; Hallam, S.J.; Delong, E.F.; Reeve, J.N., Histones in crenarchaea. J Bacteriol 2005, 187, 5482-5485.
35. Cavalier-Smith, T.; Chao, E.E., Multidomain ribosomal protein trees and the planctobacterial origin of neomura (eukaryotes, archaebacteria). Protoplasma 2020, 257, 621-753.
36. Cech, T.R., The RNA worlds in context. Cold Spring Harb Perspect Biol 2012, 4, a006742.
37. Cech, T.R., RNA World research-still evolving. RNA 2015, 21, 474-475.
38. Noller, H.F., Evolution of Protein Synthesis from an RNA World. In RNA World: From life's origin to diversity in gene regulation, Atkins, J.F.; Gesteland, R.F.; Cech, T.R., Eds. Cold Spring Harbor Laboratory Press: Cold Spring Harbor Laboratory, 2011; pp 141-154.
39. Smith, M.W.; Feng, D.F.; Doolittle, R.F., Evolution by acquisition: the case for horizontal gene transfers. Trends Biochem Sci 1992, 17, 489-493.
40. Doolittle, W.F., Evolution: Two Domains of Life or Three? Curr. Biol. 2020, 30, R177-R179.
41. Williams, T.A.; Cox, C.J.; Foster, P.G.; Szöllősi, G.J.; Embley, T.M., Phylogenomics provides robust support for a two-domains tree of life. Nature ecology & evolution 2020, 4, 138-147.
42. Gray, M.W.; Doolittle, W.F., Has the endosymbiont hypothesis been proven? Microbiol Rev 1982, 46, 1-42.
43. Margulis, L., Origin of eukaryotic cells; evidence and research implications for a theory of the origin and evolution of microbial, plant, and animal cells on the Precambrian earth. Yale University Press: New Haven, 1970; p 349.
44. Hurt, E.C.; van Loon, A.P.G.M., How proteins find mitochondria and intramitochondrial compartments. Cell 1986, 11, 204-207.
45. Gray, M.W., The evolutionary origins of organelles. Trends Genet 1989, 5, 294-299.
46. Gray, M.W., Origin and evolution of mitochondrial DNA. Annu Rev Cell Biol 1989, 5, 25-50.
47. Gray, M.W., The endosymbiont hypothesis revisited. Int Rev Cytol 1992, 141, 233-357.
48. Gray, M.W., Origin and evolution of organelle genomes. Curr Opin Genet Dev 1993, 3, 884-890.
49. Bonen, L.; Doolittle, W.F., On the prokaryotic nature of red algal chloroplasts. Proc. Natl. Acad. Sci. U S A 1975, 72, 2310-2314.
50. Bonen, L.; Doolittle, W.F.; Fox, G.E., Cyanobacterial evolution: Results of 16S ribosomal ribonucleic acid sequence analysis. Can. J. Biochem. 1979, 57, 879-888.
51. Gellissen, G.; Michaelis, G., Gene transfer. Mitochondria to nucleus. Ann N Y Acad Sci 1987, 503, 391-401.
52. Adams, K.L.; Palmer, J.D., Evolution of mitochondrial gene content: gene loss and transfer to the nucleus. Mol Phylogenet Evol 2003, 29, 380-395.
53. Dagan, T.; Martin, W., The tree of one percent. Genome Biol 2006, 7, 118.
54. Brennicke, A.; Grohmann, L.; Hiesel, R.; Knoop, V.; Schuster, W., The mitochondrial genome on its way to the nucleus: different stages of gene transfer in higher plants. FEBS Lett 1993, 325, 140-145.
55. Figueroa, P.; Gómez, I.; Holuigue, L.; Araya, A.; Jordana, X., Transfer of rps14 from the mitochondrion to the nucleus in maize implied integration within a gene encoding the iron-sulphur subunit of succinate dehydrogenase and expression by alternative splicing. Plant J 1999, 18, 601-609.
56. Kadowaki, K.; Kubo, N.; Ozawa, K.; Hirai, A., Targeting presequence acquisition after mitochondrial gene transfer to the nucleus occurs by duplication of existing targeting signals. Embo J 1996, 15, 6652-6661.
57. Nugent, J.M.; Palmer, J.D., RNA-mediated transfer of the gene coxII from the mitochondrion to the nucleus during flowering plant evolution. Cell 1991, 66, 473-481.
58. Kan, S.L.; Shen, T.T.; Ran, J.H.; Wang, X.Q., Both Conifer II and Gnetales are characterized by a high frequency of ancient mitochondrial gene transfer to the nuclear genome. BMC Biol 2021, 19, 146.
59. Andersson, S.G.; Zomorodipour, A.; Andersson, J.O.; Sicheritz-Ponten, T.; Alsmark, U.C.; Podowski, R.M.; Naslund, A.K.; Eriksson, A.S.; Winkler, H.H.; Kurland, C.G., The genome sequence of Rickettsia prowazekii and the origin of mitochondria. Nature 1998, 396, 133-140.
60. Sicheritz-Ponten, T.; Kurland, C.G.; Andersson, S.G., A phylogenetic analysis of the cytochrome b and cytochrome c oxidase I genes supports an origin of mitochondria from within the Rickettsiaceae. Biochim Biophys Acta 1998, 1365, 545-551.
61. Zaremba-Niedzwiedzka, K.; Caceres, E.F.; Saw, J.H.; Backstrom, D.; Juzokaite, L.; Vancaester, E.; Seitz, K.W.; Anantharaman, K.; Starnawski, P.; Kjeldsen, K.U., et al., Asgard archaea illuminate the origin of eukaryotic cellular complexity. Nature 2017, 541, 353-358.
62. Golyshina, O.V.; Pivovarova, T.A.; Karavaiko, G.I.; Kondratéva, T.F.; Moore, E.R.; Abraham, W.R.; Lünsdorf, H.; Timmis, K.N.; Yakimov, M.M.; Golyshin, P.N., Ferroplasma acidiphilum gen. nov., sp. nov., an acidophilic, autotrophic, ferrous-iron-oxidizing, cell-wall-lacking, mesophilic member of the Ferroplasmaceae fam. nov., comprising a distinct lineage of the Archaea. Int J Syst Evol Microbiol 2000, 50 Pt 3, 997-1006.
63. Krupovic, M.; Dolja, V.V.; Koonin, E.V., The LUCA and its complex virome. Nat Rev Microbiol 2020, 18, 661-670.
64. Nasir, A.; Kim, K.M.; Caetano-Anolles, G., Giant viruses coexisted with the cellular ancestors and represent a distinct supergroup along with superkingdoms Archaea, Bacteria and Eukarya. BMC Evol Biol 2012, 12, 156.