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Research in Microbiology 162 (2011) 77e91
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A new fusion hypothesis for the origin of Eukarya: better than previous
ones, but probably also wrong
Patrick Forterre a,b,*
b
a
Institut Pasteur, 25 rue du Docteur Roux, 75015 Paris, France
Institut de Ge´ne´tique et Microbiologie, Univ. Paris-Sud - CNRS UMR 8621, 91405 Orsay Cedex, France
Available online 27 October 2010
Abstract
I discuss here the possibility that Eukarya originated from the engulfment of a thaumarchaeon by a PCV (Planctomycetes, Verrucomicrobia,
Chlamydiae) bacterium, followed by invasions of NCLDV and retroviruses. The thaumarchaeon provided both informational and operational
proteins (actins, ESCRT proteins), including some essential proteins absent in other archaeal phyla (Topo IB), whereas the PVC bacterium
provided phospholipids, tubulin and the membrane coat proteins required for the formation of the nucleus. Viral invasions introduced many
proteins that are specific to modern Eukarya and produced an arms race that favoured the evolution of Eukarya toward increasing complexity.
This scenario is the best possible fusion hypothesis that can be presently proposed. However, it still requires several ad hoc assumptions to
explain the origin of the nucleus and the distribution pattern of archaeal and bacterial traits in modern Eukarya. Furthermore, it still fails to
explain convincingly the origin of eukaryal viruses and the existence of three distinct lineages of ribosomes. I conclude that Eukarya and their
viruses more probably evolved from a specific lineage, according to the three domains scenario originally proposed by Carl Woese.
Ó 2010 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.
Keywords: Archaea; Thaumarchaea; Planctomycetes; NCLDV; Retrovirus; Origin of eukaryotes
1. Introduction
The prokaryote/eukaryote dichotomy, set up in 1962 by
Stanier and Van Niel (Stanier and Van Niel, 1962), based on cell
biology, has been the dominant paradigm in biology for the last
50 years. It remains today accepted by most biologists despite
the fundamental work of Carl Woese and colleagues who
demonstrated in 1977, based on molecular biology, the existence
of three (and not two) major evolutionary cellular lineages,
Archaea, Bacteria and Eukarya (Woese and Fox, 1977; Woese
et al., 1990). The procaryote paradigm indeed satisfies the
traditional idea that evolution always progresses from simple
organisms to complex organisms (see a critique of this idea in
Forterre and Philippe, 1999). It is inherent in the terms
prokaryote/eukaryote themselves, since « pro » karyotes implies
* Institut Pasteur, 25 rue du Docteur Roux, 75015 Paris, France.
Tel.: þ33 169157489.
E-mail address: [email protected].
before « eu » karyotes (true nucleus) (Forterre, 1995; Pace,
2009). I suggested in 1992 to replace the terms prokaryote and
eukaryote by neutral ones (evolutionary speaking) such as
akaryote and synkaryotes (without and with nucleus) to designate the two types of known cellular organization (Forterre,
2002), but this proposal was completely ignored. Another
reason explaining the resistance of the prokaryote paradigm is
that we (human beings) like dichotomies, so the prokaryote/
eukaryote dichotomy is regularly revived, like the phoenix, even
if it is contradicted by facts.
In recent years, the prokaryote/eukaryote dichotomy has
been on the agenda again under a new appearance, i.e. “fusion”
hypotheses in which Eukarya originated from the association
of an archaeon and a bacterium (see discussions in Poole and
Penny, 2007; Gribaldo et al., 2010; Cavalier-Smith, 2010;
Koonin, 2010a,b, Forterre, 2010b). These hypotheses are
based on the observation that Eukarya and Archaea share
similar informational mechanisms (translation, transcription,
replication) very distinct (sometimes non homologous) from
0923-2508/$ - see front matter Ó 2010 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.resmic.2010.10.005
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P. Forterre / Research in Microbiology 162 (2011) 77e91
those of Bacteria, whereas Eukarya and Bacteria share a similar
type of membrane phospholipid, very distinct from those of
Archaea (see below). Bacteria and Eukarya also exclusively
share many proteins (often involved in so-called operational
mechanisms) whose origin is disputed. Some authors suppose
that these proteins have been all introduced in Eukarya either
by the alpha-proteobacterium at the origin of mitochondria or
by other bacteria used as preys by ancient Eukarya. In contrast,
proponents of “fusion” hypotheses argue that these proteins
testify for the existence of a specific bacterium at the origin of
Eukarya, whose proteome was supplemented by the informational proteins of an archaeon.
The fusions scenarios that have been proposed by different
authors involve various types of associations, symbioses,
syntrophy, etc. often justified by metabolic considerations to
explain why the two partners first of all associated. I will put
all these types of combinations here under the broad term
“fusion” for the sake of simplicity. Proponents of fusion
scenarios for the origin of Eukarya usually still use the old
name archaebacteria, instead of Archaea, thus perpetuating the
confusion between Bacteria and prokaryotes. In that scheme,
Bacteria (as a general term encompassing eubacteria and
archaebacteria) regain the primitive position that they have
occupied in the tree of life from the XIX century (the
Haeckel’s monera). In fusion scenarios, the three primary
domains of Woese and colleagues are thus replaced by two
primary “prokaryotic” domains, and “eukaryotes” therefore
emerged once more after prokaryotes, as a derived secondary
domain (see discussion in Gribaldo et al., 2010).
Proponents of fusion scenarios on the origin of Eukarya
often play of the confusion between Eukarya in general
(including those which lived before the acquisition of mitochondria) and modern Eukarya. The latter indeed originated
after Bacteria, since their last common ancestor already had
bona fide mitochondria. However, in the three domains
scenario, the primordial eukaryal lineage (urkaryote sensu
Woese and Fox, 1977) does not correspond to modern
Eukarya, but to a lineage of ancient Eukarya that led to LECA
(the Last Eukaryal Common Ancestor) (Fig. 1A). This lineage
might be as ancient as those that led to Bacteria and Archaea.
In this scenario, FECA (the First Eukaryal Common Ancestor)
might have even predated the divergence of the various phyla
of modern archaea and bacteria (Fig. 1A). In contrast, in
fusion scenarios, FECA only appeared after the diversification
of the archaeal and bacterial domains (Fig. 1B). Indeed, with
few exceptions, proponents of fusion scenarios for the origin
of Eukarya have always proposed that the two partners that
associate were close relatives of modern Bacteria and Archaea.
Historically, various archaeal and bacterial partners have
been proposed by different authors to sell convincingly their
particular fusion hypotheses. Lynn Margulis, suggested in the
eighties that the archaeal partner was a relative of Thermoplasma acidophilum (Margulis and Bermudes, 1985),
a member of the archaeal phylum Euryarchaeota, because T.
acidophilum is a wall-less microorganism in which “eukaryotic-like histones” were described (biochemically) in the
seventies (Stein and Searcy, 1978). In her hypothesis, the
bacterial partner was a Spirochete because these bacteria
harbour microtubule-like structures reminiscent of the eukaryal
cytoskeleton (Margulis et al., 1979). Later on, Rivera and Lake
suggested an archaeon of the phylum Crenarchaeota for the
archaeal partner, because Crenarchaeota (called Eocytes by
Lake) were sister groups of Eukarya in their phylogenetic
analyses (Rivera and Lake, 1992, 2004). More recently, in the
“ring of life” scenario, they identified the bacterial partner as
a relative of Proteobacteria and/or Cyanobacteria (Rivera and
Lake, 2004). In the so-called, “hydrogen hypothesis”, Martin
and Muller suggested an unusual version of fusion hypotheses
in which the bacterial partner corresponds to the alpha-proteobacterium at the origin of mitochondria (Fig. 1B, arrow b).
In their scenario, the “archaebacterium” is an unspecified
autotrophic hydrogen-dependent anaerobe (Martin and Müller,
1998). One of the more detailed and original fusion hypothesis
is the “syntrophic hypothesis” proposed by Moreira and Lopez-
Fig. 1. Two alternative scenarios for the origin and evolution of modern domains. A: Scenario based on the three domains concept (dotted lines symbolize an
unrooted tree), B: Fusion scenario based on the prokaryote/eukaryote dichotomy. The green arrows represent the origin of the mitochondrion from an alphaproteobacterium. FECA: First Eukaryotic (specific) Common Ancestor, LECA: Last Eukaryotic Common Ancestor, FME: First Mitochondrial Eukaryote.
P. Forterre / Research in Microbiology 162 (2011) 77e91
Garcia (1998). These authors suggested a mesophilic methanogen (more precisely a Methanobacteriale) for the archaeal
partner, and a delta-proteobacterium (more precisely an
ancestral sulfate-reducing myxobacterium) for the bacterial
one, their complementary physiology being supposed to
explain their association based on a metabolic symbiosis
(interspecies hydrogen transfer). They also justified the choice
of a methanogen on molecular basis, i.e. the presence in these
mesophilic archaea of eukaryal histone homologues (lacking in
Crenarchaea/Eocytes). Similarly, a myxobacterium was
proposed as bacterial partner because these social and complex
bacteria were supposed to have provided the primordial genes
for the social and developmental behaviour of later Eukarya,
and because Myxobacteria were supposed to be at that time the
only “prokaryotes” encoding serine/threonine protein kinases
or small G-proteins (supposed to be eukaryotic hallmaks).
Later on, the idea of an association between a methanogenic
archaeon and a bacterium was also endorsed by Bell. This
author combined the scenarios of Moreira and Lopez-Garcia
with the hydrogen hypothesis of Martin and Muller (but
removing the myxobacterium) and introduces in the story
a giant Pox-like virus infecting the methanogenic archaeon. In
this me´nage à trois, the alpha-proteobacterium was transformed
into a mitochondrion, whereas the giant virus became the
nucleus of modern Eukarya (Bell, 2001).
Detailed fusion hypotheses for the origin of Eukarya made
during the last years are now open for falsification (or at least
criticism) following new discoveries in archaeal and/or bacterial
physiology, molecular biology and genomics. These discoveries
often make obsolete the reasons that initially justified the choice
of a particular partner of the fusion. For instance, the case for
a Thermoplasma-like archaeon at the origin of the eukaryotic
nucleus became much weaker when it turned out that the
“eukaryal-like histones” of Thermoplasmatales are not homologous to eukaryal histones but to the bacterial HU protein
(DeLange et al., 1981; Sandman et al. 1990). Similarly, the idea
79
that Eukarya partly derived from methanogens because the
presence of eukaryal-like histones in these mesophilic archaea
became obsolete when similar histones were discovered in
mesophilic thaumarchaea (Cubonová et al., 2005, BrochierArmanet et al., 2008a,b). The case for a spirochete or a deltaproteobacterium at the origin of Eukarya also became weak once
the sequencing of their genomes failed to reveal specific affinities
with Eukarya and after the discovery of protein kinases and Gproteins in many groups of Archaea and Bacteria (Dong et al.,
2007; Tyagi et al., 2010).
The choice of two specific partners in fusion hypotheses has
always been dependant of data available at a given time in
terms of microbial diversity, physiology and molecular biology.
Interestingly, recent discoveries have revealed novel eukaryal
features in members of the bacterial PVC superphylum, that
includes Planctomycetes, Verrucomicrobia, Chlamydiae, Lentisphaera and Poribacteria (Wagner and Horn, 2006), and in
members of the recently recognized archaeal phylum Thaumarchaeota (whose mesophilic members were formerly
assimilated to Crenarchaeota) (Brochier-Armanet et al.,
2008a). Proteins similar (probably homologous) to eukaryal
membrane coat (MC) proteins have been discovered in PVC
bacteria (Santarella et al., 2010), whereas several eukaryal-like
features not found in other archaea have been detected in
Thaumarchaeota (Brochier-Armanet et al., 2008a,b; Spang
et al., 2010). PVC bacteria and Thaumarchaeota have not
been previously suggested as potential fusion partners for the
origin of Eukarya. Before someone else thus proposes a new
fusion scenario involving these two groups, I decided to do it
myself. In this new scenario, FECA originated from the
engulfment of a thaumarchaeon (the symbiont) by a PVC
bacterium (Fig. 2). This fusion was followed by the massive
invasion of the FECA descendants by several novel lineages of
viruses, such as NCLDV (Nucleo-Cytoplasmic Large DNA
Viruses) and retroviruses (Fig. 2). Integration of viral genomes
into the genomes of FECA descendants introduced new
Fig. 2. The PTV fusion hypothesis based on the engulfment of a thaumarchaeon by a PVC bacterium followed by viral invasions. Bacterial and archaeal
membranes, cytoplasmic and nuclear components (including circular chromosome) are in green and purple, respectively. Eukaryal cytoplasmic and nuclear
components are in grey to symbolize differences with their archaeal ancestors. Eukaryal chromosomes (linear) are in orange. Abbreviations are as in Fig. 1. ICM:
Intracytoplasmic membrane. NE: nuclear envelope. PVC: Planctomycetes, Verrucomicrobia, Chlamydiae superphylum. For simplicity the reticulum endoplasmic
membrane deriving from the ICM of the PVC bacterium has not been indicated.
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P. Forterre / Research in Microbiology 162 (2011) 77e91
proteins (and protein folds) into ancient eukaryal lineages,
explaining the presence in modern Eukarya of many proteins
without homologues in Archaea and Bacteria. Furthermore,
intense competition between viruses and ancient Eukarya
produced an arms race that favoured evolution toward
increasing complexity from FECA to LECA. For instance, the
invasion by retroviruses might have triggered the transformation of circular chromosomes into linear ones and initiated the succession of events at the origin of sex. This new
hypothesis will be dubbed the PTV scenario thereafter (for
PVC bacteria eThaumarchaea - Virus).
I choose a bacterial host because it readily explains why the
phospholipids of eukaryal cell membranes (the cytoplasmic
membrane, the endoplasmic reticulum and the nuclear
membrane) are made of bacterial type phospholipids (esters
of sn-glycerol-3-phosphate and fatty acid) (green in Fig. 2).
Hypotheses in which the host is an archaeon should explain
why archaeal phospholipids (ethers of sn-glycerol-1-phosphate and isoprenol) were replaced by bacterial ones in all
these membranes. This is really impossible without spurious
ad hoc assumptions. Although unlikely (see below), it seems
easier to postulate that the internal membrane of the endosymbiont (the archaeon) disappeared after the fusion, than to
assume (if the symbiont was a bacterium) that the bacterial
lipids of the symbiont replaced those of the host!
I will thereafter explain why, although this new proposal is
probably the best possible fusion scenario that one can
imagine with our present knowledge, I still don’t believe in it.
Finally, I will discuss how some novel aspects of this scenario
(in particular the intervention of viruses) could be nevertheless useful for imaging how Eukarya originated and evolved
from the Last Universal Cellular Ancestor (LUCA) in the
framework of the three primary domains concept of Carl
Woese.
2. A new fusion scenario: a PVC bacterium as the host
and a thaumarchaeon as the symbiont
2.1. Why a PVC bacterium
The PTV scenario posits that the host bacterium at the origin
of Eukarya was evolutionarily related to modern bacteria of
the superphylum PVC because genes encoding structural
analogues of eukaryal MC proteins, such as clathrins and
nucleoporins, have been recently detected in the genomes of
several Planctomycetes, Verrucomicrobia and Lentisphaera
(Santarella et al., 2010). These proteins exhibit a unique
structure formed by an amino-terminal beta-propeller domain
followed by a carboxy-terminal stacked pairs of alpha-helices.
The respective length of the two domains, as well as the
respective number of beta strands and alpha helices in each
domains are strikingly similar in PVC bacteria and Eukarya,
strongly suggesting that MC proteins of PVC bacteria and
Eukarya are homologous (5). Similar proteins have never been
found in Archaea or in Bacteria other than PVC bacteria. In
Eukarya, MC proteins are involved in the formation of the
endoplasmic reticulum (e.g. clatherins and related proteins)
and of the nuclear pores (e.g. nucleoporins). Strikingly,
PVC bacteria encoding eukaryal-like MC proteins exhibit an
intracytoplasmic membrane (ICM) mimicking the endoplasmic
reticulum of Eukarya (Fuerst, 2005; Lee et al., 2009; Santarella
et al., 2010). In Eukarya, the endoplasmic reticulum membrane
is recruited to form the double membrane that surrounds the
chromosomes (the nuclear envelope). Similarly, in the Planctomycete Gemmata obscuriglobus, a synkaryotic bacterium, the
ICM is recruited to form a double membrane that surrounds the
bacterial nucleoid (Fuerst, 2005). Recently, it has been shown
that G. obscuriglobus can perform endocytosis, a property that
was until now thought to be restricted to Eukarya, and one of its
MC proteins seems to be involved in that process (Lonhienne
et al., 2010). PVC Bacteria thus appear the ideal hosts for
a fusion hypothesis, since they could explain the emergence in
Eukarya of both the endoplasmic reticulum and the nuclear
envelope.
Beside the presence of eukaryal-like MC proteins, PVC
bacteria exhibit several additional features that favour them as
putative hosts in fusion hypotheses for the origin of Eukarya. In
particular, synkaryotic PVC bacteria (G. obscuriglobus and
Poribacteria) are among the few bacteria in which sterol
biosynthesis has been documented (Pearson et al., 2003; Siegl
et al., in press). Otherwise, sterol biosynthesis is almost
exclusively restricted to Eukarya. Plancomycetes are quite large
cells that divide by budding, a feature that reminds us of
Eukarya (in fact, Plancomycetes were once confused with
fungi, hence their name) (Fuerts, 1995). Similar to Eukarya,
some PVC bacteria also lack peptidoglycan and the bacterial
cell division protein FtsZ (Pilhofer et al., 2008). This favours
PVC bacteria over other bacteria as potential hosts able to
engulf small cellular symbionts. All known modern PVC
bacteria have cell envelope and cannot perform phagocytosis.
However, one can imagine that, building on their ability to
perform endocytosis, some ancient or not yet discovered wallless PVC bacteria (L-forms) have used their MC protein to
evolved the ability to engulf small preys. Importantly, some
Planctomycetales relatives (anammox bacteria) contain an
intracellular organelle, the anammoxosome, surrounded by
a membrane formed by atypical lipids (ladderanes) (Jetten et al.,
2009). The membrane of the anammoxosome contains an FATP synthase, mimicking the cytoplasmic membrane of a bona
fide organisms (Van Niftrik et al., 2010). The origin of the
anammoxosome is unknown since this organelle lacks DNA.
However, in the framework of the present hypothesis, it is
tempting to suggest that some ancestor of anammox bacteria
have had the capacity to serve as host for other microbes.
Finally, closely related homologues of the two eukaryal
tubulins (alpha and beta) have been found in bacteria of the
genus Prosthecobacter, which are members of Verrucomicrobia (Jenkins et al., 2002). Interestingly, the alpha and beta
tubulins of Prosthecobacter do not branch within Eukarya in
a tubulin phylogenetic tree, but as outgroups of eukaryal alpha
and beta eukaryotic tubulins, respectively, arguing against
a lateral gene transfer from a modern Eukarya to this group of
PVC bacteria (Jenkins et al., 2002). In the PTV hypothesis, the
two tubulin genes originated first in PVC bacteria and were
P. Forterre / Research in Microbiology 162 (2011) 77e91
later on transferred to Eukarya via the bacterial partner of the
fusion.
In summary, in the PTV scenario, the PVC bacterium
provided to the future Eukarya not only membrane phospholipids (as in other fusion scenarios) but also the ICM
membrane, tubulin and MC proteins (precursors of the cytoskeleton, nucleoproteins and of the nuclear pore complex), as
well as the ability to synthesize sterols, a universal component
of eukaryal membranes (Table 1).
2.2. Why a thaumarchaeon as the symbiont?
In the PTV scenario, the archaeal symbiont was a thaumarchaeon because these archaea exhibit a wider range of
eukaryal features than archaea from the two other phyla, Euryarchaeota and Crenarchaeota (Brochier-Armanet et al.,
2008a,b; Spang et al., 2010) (Table 1). Hence, whereas the
large RNA polymerase A subunit is splitted in Crenarchaea and
Euryarchaea, it corresponds to a single polypeptide in Thaumarchaea, as in Eukarya and Bacteria (Kwapisz et al., 2008).
Thaumarchaea are also the only Archaea to harbour a type IB
DNA topoisomerase, the enzyme responsible in Eukarya to
relax positive supeturns that accumulate during DNA replication and transcription (Brochier-Armanet et al., 2008b). They
are also the only Archaea that contain the two single-stranded
DNA binding proteins (SSB) found in Eukarya, the protein
RPA, previously thought to be restricted to Euryarchaea, and
the small SSB recently found in human (hSSB), previously
thought to be restricted to Crenarchaeota (unpublished observation). Thaumarchaea harbour eukaryal-like histones, otherwise mainly present in Euryarchaeota (Cubonová et al., 2005),
and the two eukaryotic-like “cell division” proteins (CdvB/Snf7
Table 1
Molecular features introduced in Eukarya by the thaumarchaeon, the PVC
bacterium and viruses, respectively, in the framework of the PTV scenario.
Molecular features either present in Thaumarchaea but not in other archaeal
phyla or present in some PVC bacteria but not in other bacterial phyla are in
bold.
Thaumarchaeon
PVC bacterium
Viruses
Translation machinery
(ribosomes)
Transcription machinery
Non split RpoA
Transcriptional
regulators
mRNA capping
New basal transcription
factors
Spliceosomes (?)
New DNA polymerases
Topo IIA
Linear chromosomes
Telomers, Centromers
Sex
Replication machinery,
histones
RPA D cren SSB
Topo IB
Bacterial lipids
ESCRT proteins
Membrane Coat
V-ATP ase, Sec system, Proteins
SRP
Intracytoplasmic
membrane (ICM)
Sterols
Actin
Tubulin
New proteins to manipulate
membranes previously
involved in formation of
viral factories
New cytoskeletal proteins
of low-complexity
sequence composition
81
and CdvC/Vps4), otherwise mainly present in Crenarchaeota
(for review, see Makarova et al., 2010). In Eukarya, the proteins
CdvB/Snf7 and CdvC/Vps4 belong to the ESCRT complex
(Endosomal Sorting Complex Required for Transport) that is
involved in vesicles formation and cell division. Importantly, the
archaeal proteins seem to be involved in similar mechanisms
(Samson and Bell, 2009; Ettema and Bernander, 2009).
In the PTV scenario, I will suppose that the thaumarchaeon
at the origin of Eukarya also contained the archaeal actin protein
(Arch-actin) recently detected in Thermoproteales (members of
the phylum Crenarchaeaota) and in the only Korarchaeon
whose genome has been sequenced, "Candidatus Korarchaeum
cryptofilum" (Yutin et al., 2009). The Arch-actin is much more
similar to eukaryal actin than other actin homologues previously found in Bacteria or Archaea, MreB and FtsA, respectively (for review, see Makarova et al., 2010). Although the
three presently Thaumarchaeota whose genomes have been
sequenced lack the Arch-actin gene, genome context analysis
suggests that this gene was present in the Last Archaeal
Common Ancestor (LACA) and was later on lost in the branch
leading to the three Thaumarchaea whose genomes have been
sequenced (Makarova et al., 2010). In the PTV scenario, a major
consequence of the fusion was thus to bring for the first time
actin (from the thaumarchaeon) and tubulin (from the PVC
bacterium) in the same organism and later on (once the archaeal
membrane has disappeared) in the same cytoplasm.
In previously proposed fusion hypotheses, one has to assume
a dramatic acceleration of the rate of protein evolution after
the fusion to explain why are tubulin and actin now so different
in Eukarya from their distant homologues in Bacteria and
Archaea, FtsZ and MreB proteins, respectively (Doolittle, 1995;
Erickson, 2007). This is especially difficult to admit if the
bacterial partner was the alpha-proteobacterium at the origin of
the mitochondrion (as in the hypothesis of Martin and Muller).
Why did the FtsZ of the alpha-proteobacterium evolved beyond
recognition (at the sequence level) from its bacterial ancestor,
whereas many other proteins from the same alpha-proteobacterium remain so similar in Bacteria and mitochondria. To
take a single example, the mitochodrial protein Qri7 branches as
sister group of the alpha-proteobacterial YgjD (its ancestor) in
phylogenetic trees, and the mitochondrial protein can complement a YgjD bacterial mutant (Oberto et al., 2009). The
dramatic transformation of FtsZ and MreB proteins into tubulin
and actin was therefore a major drawback of previous fusion
scenarios. In contrast, in the PTV scenario, eukaryal tubulin and
actin did not originate from bacterial FtsZ and MreB proteins,
but directly from archaeal actin and bacterial tubulin, with
sequences already very similar to those of modern Eukarya.
In addition to exhibit a unique combination of eukaryoticlike features not found in other archaeal phylums, Thaumarchaea are also good candidates to be the archaeal symbiont
in a fusion scenario because they are abundant in marine,
lacustre and terrestrial mesophilic environments where they
coexist with bacteria, including PVC bacteria. For instance,
Cenarchaeum symbiosum (a thaumarchaeon) and Poribacteria
(PVC bacteria) live as intracellular symbionts within sponges.
Of course, one cannot argue that a bacterium and an archaeon
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P. Forterre / Research in Microbiology 162 (2011) 77e91
meet in a sponge to produce the ancestor of sponges! Nevertheless, many possibilities remain open.
I will not try to propose here a specific type of metabolism
complementation to justify the PTV scenario. Indeed, both
Thaumarchaea and PVC bacteria are very diverse but poorly
studied groups that probably include many types of metabolisms. For instance, the capacity of some thaumarchaea to
oxidize ammonium has only been discovered five years ago
(Treusch et al., 2005). I thus assume that Thaumarchaea and
PVC Bacteria with complementary metabolisms will be
discovered soon or later to justify their association in the PTV
scenario. It will then be possible to imagine the specific type
of “fusion” suitable to associate these complementary partners
and the type of environment in which they meet each other.
In summary, in the PTV scenario, the thaumarchaeon
provided to the future Eukarya all proteins and machineries that
remain highly similar today between Archaea and modern
Eukarya. These proteins and mechanisms (e.g. ribosomes, DNA
replication and transcription apparatus) are typically bacterial
in PVC bacteria, indicating that Eukarya cannot be derived
directly from a PVC bacterium, even if the latter is a synkaryote.
Proteins provided by the thaumarchaeon included not only
informational proteins but also operational proteins, such as
some precursors of the modern eukaryal cytoskeleton (actin)
and vesicle trafficking systems (ESCRT proteins) (Table 1).
2.3. Origins of mitochondria
In the PTV scenario, the endosymbiotic event that produced
mitochondria occurred obviously after the fusion event that
produced the FECA (Fig. 2). At first sight, fusion hypotheses in
which the bacterial partner became the mitochondrion (arrow
b in Fig. 1B) seems more parsimonious because they have no
need to postulate the existence of ancient Eukarya without
mitochondria (Archezoa, sensu Cavalier-Smith, 1989). For
some authors, this explains the failure to discover such Archezoa today (living fossils). However, it is crystal clear that
ancient Eukarya should have existed, whatever the scenario we
prefer for the origin of Eukarya. For instance, reconstruction of
the mitochondrion present in the LECA clearly indicates that
mitochondria have evolved a long way between the alphaproteobacterium present in the first mitochondrial Eukarya
(FME) and the mitochondrion present in LECA. Indeed, phylogenomic analysis shown that the mitochondrial ribosome of
LECA has already gained 19 novel proteins that were not
present in its bacterial ancestor (Desmond et al., 2011).
Therefore, even if the FECA and the FME were the same
organism, one has to postulate the existence of many intermediate forms of ancient Eukarya which had mitochondrial ribosomes with protein compositions intermediate between those of
the FECA/FME and the LECA (Fig. 1B). Furthermore, in any
scenario, the emergence of specific eukaryal features already
present in LECA such as the nucleus, the cytoskeleton, spliceosomes, or the mitotic apparatus, should have spanned a long
evolutionary period (Gribaldo et al., 2010). For example, it is
quite clear that meiosis evolved from mitosis through a complex
evolutionary pathway involving several successive steps
(Wilkins and Holliday, 2009). Ancient Eukarya (urkaryotes,
sensu Woese and Fox, 1977) have thus certainly existed for
a long period of time (co-existing with Archaea and Bacteria)
Fig. 3. The three possible (but unlikely) hypotheses for the origin of the eukaryal nuclear envelope in the PTV scenario. A: the eukaryal nuclear envelope derived
directly from the nucleus of a synkaryotic PVC bacterium, such as Gemmata obscuriglobus. B, the eukaryal nuclear envelope derived from the ICM of an akaryotic
PVC bacterium. C: the eukaryal nuclear envelope derived directly from the membrane of the thaumarchaeon. Membranes with bacterial type and archaeal type
phospholipids are in green and purple, respectively. Bacterial and archaeal cytoplasmic components and chromosomes are in green and purple, respectively. Colour
shift from green to purple symbolizes the replacement of bacterial components (e.g. ribosomes) by archaeal ones. In the three scenarios, FECA is transformed into
a proto-eukarya before the mitochondrial endosymbiosis, and not yet remodelled by extensive viral invasion. For simplicity the reticulum endoplasmic membrane
deriving from the ICM of the PVC bacterium (A, B) or from the modified envelope of the thaumarchaeon (C) has not been indicated. Abbreviations are as in Figs. 1
and 2.
P. Forterre / Research in Microbiology 162 (2011) 77e91
and have diversified in many lineages that have all disappeared
today (much like pre-Homo sapiens during Homo evolution)
(Fig. 3). The existence of extinct lineages is indeed a consequence of biological evolution already discussed by Darwin in
the “origin of species”.
3. The viral hypothesis for the origin of eukaryal
complexity
The existence of numerous complex and specific molecular
features in Eukarya (either specific protein families or specific
molecular mechanisms) has always been a major challenge to
proponents of fusion hypotheses (Koonin, 2010). In the most
recent reconstitution of the putative proteome of LECA (performed after the sequencing of the protist Naegleria gruberi)
Fritz-Laylin and colleagues concluded that at least 40% of
proteins in LECA e 1420 - were “eukaryotic inventions” (they
have no detectable archaeal or bacterial homologues) (FritzLaylin et al., 2010). In the PTV scenario, some eukaryal
specific proteins might correspond to thaumarchaeal or PVC
bacterial proteins that have been lost in the few genomes of
PVC bacteria or Thaumarchaea presently available. As a matter
of fact, the archaeal orthologues of actin and Topo IB have been
only recently detected in modern archaeal genomes. Further
archaeal or bacterial homologues of eukaryal specific proteins
will be also probably detected in the future at the structural
level, as it was recently the case for MC proteins of PVC
bacteria. However, it seems unlikely that “prokaryotic” homologues will be finally found for all eukaryal specific protein
families and protein folds. Indeed, even before the sequencing
of N. gruberi, Kurland and colleagues identified in the universe
of protein folds 244 fold superfamilies that are specific to
Eukarya, versus 16 and 49 that are specific for either Archaea or
Bacteria, respectively (Kurland et al., 2007). Furthermore, it
is unlikely that we will find in the future eukaryotic specific
mechanisms, such as mRNA capping and splicing, telomers,
meiotic sex, mid-bodies and so on, in some PVC bacteria or
Thaumarchaeota. The creation of all these eukaryal novelties
has been a major puzzle for all fusion hypotheses. The only
possibility to bypass this problem is to suggest that the fusion
triggered an explosion of creativity, leading to a dramatic
increase in complexity of the resulting entity. However, there is
no known example in current biology of such situation. Usually,
a combination of organisms led to the reductive evolution of
one of them, but never to an explosion of creativity and drastic
modifications in the molecular make up of the other. For
instance, the “fusion” (endosymbiosis) between Eukarya and
Cyanobacteria did not change the basic molecular biology of
Eukarya, although it triggered the emergence of a new eukaryal
division (Viridiplantae).
In the framework of the PTV scenario, I suggest here a new
hypothesis that could help to solve this conundrum. I propose
that eukaryal specific protein families and mechanisms originated as the consequence of several massive viral invasions
that pecifically shaped the evolution of the eukaryal lineage
from FECA to LECA. In this hypothesis, multiple rounds of
integration of viral genomes into the genomes of ancient
83
Eukarya introduced new proteins (and protein folds) into the
descendants of FECA, whereas the intense competition
between viruses and ancient Eukarya produced an arms race
that favoured evolution toward increasing complexity from
FECA to LECA.
What makes plausible the idea that repeated viral invasions
introduced in ancestral Eukarya new protein families without
archaeal or bacterial homologues is that most proteins encoded by viral genomes today have no cellular homologues in
protein databases (up to 100% in the case of some archaeal
viruses) (Prangishvili et al., 2006b; Yin and Fischer, 2008;
Boyer et al., 2010) and that most viral proteins whose
structure has been solved harbour unique folds (Bamford
et al., 2005; Saini and Fisher, 2007, for a recent case
studies, see Keller et al., 2009). Genes are indeed created
continuously de novo in viral genomes within the infected
cells (virocells, sensu Forterre, 2010b) by the same processes
that create new genes in cellular genomes (gene duplication,
gene slippage, overprinting and so on) (Rancurel et al., 2009;
Forterre, 2010b).
In the PTV scenario. I suggest that ancestors of modern
NCLDV most likely played a major role in the introduction of
new protein families in Eukarya. Ogata and Claverie indeed
concluded from an analysis of proteins families specific for
various lineages of NCLDV that these viruses with large
genomes (between 150 and 1200 kb) act indeed as “an invention
factory for new genes” (Ogata and Claverie, 2007). These
viruses are very ancients, as the diverse NCLDV families
probably diverged even before LECA (Koonin and Yutin,
2010). Recently, large portions of an NCLDV genome were
found integrated in the genome of the Brown algae Ectocarpus
siliculosus, indicating that NCLDV specific genes can become
incorporated into eukaryal genomes, becoming “eukaryal
specific genes” (Cock et al., 2010). The genomes of modern
NCLDV carry a relatively high percentage of genes with
eukaryal homologues (10e20%) that are usually supposed to be
derived from eukaryal genomes (Moreira and López-Garcı́a,
2009). In fact, many of them have probably originated first in
NCLDV and were later on recruited by Eukarya (discussed in
Forterre, 2010a). Interestingly, Ogata and Claverie noticed
that NCLDV family-specific proteins are enriched in lowcomplexity sequences, resembling eukaryal proteins involved
in nucleic acid-interactions and in the architecture of the
cytoskeleton (Ogata and Claverie, 2007). Proteins with lowcomplexity sequences could originate more frequently in
viruses since mistakes such as replication slippage or abnormal
recombination probably occur more frequently in viral genomes
than in cellular genomes. I thus suggested previously that many
eukaryal proteins with low-complexity sequences involved in
interactions with nucleic acids or in cytoskeleton architecture
first originated in NCLDV and were later on introduced in
Eukarya (Forterre, 2010a). Beside NCLDV, other viruses such
as Retroviridae, Adenoviridaea, Herpesviridae, Baculoviridaea
or still other not yet discovered viral lineages (for instance
among viruses infecting protists that have been very poorly
screened) could have been contributed to the introduction of
new proteins in Eukarya.
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P. Forterre / Research in Microbiology 162 (2011) 77e91
In the PTV scenario, viral invasions not only provide
new protein families to ancient Eukarya but also induced
a continuous arms race between viruses and eukaryal cells that
dramatically altered the outcome of eukaryal evolution; this
arms race leading to the emergence of more and more complex
organisms, with both the creation of entirely novel molecular
mechanisms (e.g. mRNA capping, see below) and improvement of ancient already existing ones (e.g. transcription).
As an example, let’s see how the viral invasion might have
triggered the formation of the odd cap structures that tag in
50 eukaryotic messenger RNA (unknown in Archaea and
Bacteria). NCLDVencode homologues of the eukaryal enzymes
responsible for the formation of this structure (Shuman, 1995).
Imagine that an ancient NCLDV first introduced a simple tag,
“A”, into the 50 end of its mRNA in order to degrade selectively
cellular mRNA (the tag providing protection of mRNA of the
virus against its own nuclease). As a result, cells will be later on
selected that have aquired the ability to label their own mRNA
with the viral tag A. In response, viral mutants producing tag
of increasing complexity (AB) will then be selected. This will
later favour mutant cells having aquired this “AB” tag, leading
to the appearance of viruses with even more complex tag (ABC)
and so on. A strong argument in favour of such viral scenario
for the origin of mRNA tags is that two different types of
complex mRNA tags have now been documented in viruses
(Koonin and Moss, 2010; Ogino et al., 2010), whereas only one
of those exists in cellular mRNA. The type of scenario suggested
here to explain the origin of the complex eukaryal mRNA
capping system can be readily applied to other systems,
explaining why many molecular mechanisms become so
sophisticated in Eukarya. The spliceosome and the various
layers (mediator and so on) of the eukaryal transcription apparatus, including its regulation, are hence prime players in cellvirus molecular interactions.
If the PVC bacterium partner in the fusion was an akaryore
(with no nucleus, but simply an ICM), viruses might have also
play a major role in the emergence of the eukaryal nucleus.
Indeed, many DNA and RNA viruses that replicate in the
cytoplasm of eukaryal cells recruit the reticulum endoplasmic
membrane of their hosts (victims) to build their viral factories,
much like Eukarya recruit the endoplasmic reticulum
membrane to form their nucleus. Among them, Mimivirus (an
NCLDV) produces viral factories that are as big as the nucleus
of an Amoeba (Suzan-Monti et al., 2007). One can therefore
imagine that proteins used by one such virus to produce viral
factories in the PVC bacterium were recruited by an ancient
eukaryal cell to build its nucleus from the PVC ICM. The viral
factory is built as to protect the viral genome against the
defences of the host. Similarly, the nucleus might have been
first built to protect the cellular genome against invading
viruses (Forterre and Prangishvili, 2009b; Bandea, 2009).
A major feature that distinguishes modern Eukarya from
Archaea and Bacteria is the structure of their chromosomes.
In the PTV scenario, retroviruses have probably played
a crucial role in transforming the circular chromosome(s) of
FECA into the linear chromosome of modern eukaryal cells.
Indeed, both telomeres and centromeres appear to be related
to retroviruses. Telomerase is homologous to reverse transcriptase (Arkhipova, 2006) and it is intriguing that Penelope
retroelements containing telomeric-like sequences directly
integrate at chromosome ends (Gladyshev and Arkhipova,
2007), whereas centromeres are rich in repeated sequences
ultimately of retroviral origin (Chueh et al., 2009).
Retroviruses and derived elements might have also been
major players in the origin of sex in Eukarya, and, more
specifically, in the evolution from mitosis to meiosis. Wilkins
and Holliday (2009) have recently suggested that the first step
in the evolution of meiosis was the emergence of a mechanism
to pair homologous chromosomes. In their hypothesis, this
mechanism was selected to prevent potentially damaging
recombinations between repeated regions present in non
homologous chromosomes. If the chromosomes of ancient
Eukarya have been invaded by retroviruses, integrated retroviruses have been most likely a major source of repeated
sequences in their chromosomes. In that case, retroviruses and
retroelements might have provided the selection pressure for
the evolution of meiosis from mitosis by the mechanism
proposed by Wilkins and Holliday.
The introduction of viruses in the PTV scenario could
a priori solve another major problem (at least partly see below)
that plague all fusion hypotheses: why eukaryal proteins of
archaeal origin experienced such a dramatic acceleration of
their evolutionary rate after the fusion, such that most of them
do not branch within Archaea in phylogenetic tree, but as sister
group of Archaea (Yutin et al., 2008, see discussion in Gribaldo
et al., 2010). Proponents of fusion hypotheses usually argue that
this acceleration was induced by the fusion itself that led to
a very peculiar chimera. However, as already discussed in the
case of the mirochondrial protein Qri7 and its bacterial homologue YgjD, actual fusion between Eukarya and Bacteria did
not induce such acceleration of the rate of protein evolution. For
another example, the DNA gyrase, of bacterial origin, which is
today encoded in the nuclear genome of plants and is targeted to
both chloroplats and mitochondria, still branches within
Bacteria, as sister group of Cyanobacteria in the phylogenetic
tree of type II DNA topoisomerases (Forterre et al., 2007). This
protein has therefore evolved very slowly after the endosymbiosis event that led to the chloroplast, despite a complete
modification of its cellular environment and likely modification
and diversification of its function. Thus, evolutionary rate
acceleration is not an inevitable outcome of endosymbiotic
fusion events.
In the PTV scenario, one can imagine that most eukaryal
genes have shuttled (after the fusion) between the genomes of
ancient Eukarya and those of their invading viruses. During
their passage in viral genomes, these genes would have evolved
at a more rapid pace than their archaeal sisters that remained
most of the time confined in cellular genomes. We have recently
observed such a phenomenon in studying the evolution of the
replicative helicase MCM in Archaea (Krupovic et al., 2010).
An MCM protein of cellular origin that had transited by viral
genomes in Methanococci evolved much more rapidly than its
orthologues in other archaeal lineages. A similar scenario could
explain, for example, why some eukaryal DNA replication
P. Forterre / Research in Microbiology 162 (2011) 77e91
proteins diverged so much from their archaeal ancestors, since
DNA replication proteins appear to be frequently captured by
viruses. This is for instance the case of some eukaryal cellular
DNA polymerases that are much more related to DNA polymerases of NCLDV than to archaeal DNA polymerases
(Villarreal and DeFilippis, 2000; Filée et al., 2002).
The importance of viruses in the PTV scenario is in line
with the recent recognition of their major role in the evolution
of life on our planet (Forterre, 2005, 2006; Comeau and Krish,
2005; Forterre and Prangishvili, 2009a,b, Brüssow, 2009;
Bandea, 2009; Cortez et al., 2009; Villarreal and Witzany,
2010). In the PTV scenario, the evolution of Eukarya toward
complexity from FECA to LECA occurred by combining the
power of viral creativity (import of new proteins) and selection
pressure (arms race type of evolution) (Comeau and Krish,
2005; Forterre and Prangishvili, 2009b). Importantly, the
role of viruses in this scenario satisfies the principle of
continuity, being consistent with their roles in recent eukaryotic evolution. Viruses have been indeed essential in the
creation of novelties such as the unique replication and transcription system of mitochondria (Filée and Forterre, 2005) or
else the formation of the placenta in Mammals (Rote et al.,
2004).
4. Why the best possible fusion hypothesis is probably
wrong
The PTV scenario solves several questions raised by fusion
hypotheses in general, and proposes much more convincing
“prokaryotic” partners than previous hypotheses. However, I
don’t think that this scenario can finally save fusion hypotheses
from being fundamentally flawed, and I will discuss now why.
Firstly, despite the presence of a nucleus in some PVC
bacteria, it remains difficult to explain the origin of the eukaryal
nucleus, since all molecular mechanisms working today within
the eukaryal nucleus are much more similar to their archaeal
than to their bacterial countrparts. In the PTV scenario, the
eukaryal nucleus might be either a direct descendant of
a nucleus already present in the host PVC bacterium (a synkaryote) or it might have originated after the fusion, from the
ICM of the PVC bacterium (an akaryote) (Fig. 3A). The first
hypothesis seems a priori more parsimonious since it implies
that the “invention” of the nucleus occurred only once in life
history (in PVC bacteria) and that the fusion directly produced
a FECA with a nucleus (Fig. 3A). However, this hypothesis
is difficult to reconcile with the fact that the DNA replication
and transcription machineries, as well as the chromatin structure (histones) of modern Eukarya should have originated from
the archaeal symbiont. If the eukaryal nucleus derived directly
from the nucleus of the PVC bacterium host, one should thus
assume that all informational genes of bacterial origins were
replaced in situ (in the bacterial chromosome) by archaeal
genes encoding similar function recruited from the chromosome of the thaumarchaeon. There is no obvious selection
pressure to explain this highly unusual event. Furthermore, the
thaumarchaeon (especially its membrane) should have
completely disappeared in the process (Fig. 3). Massive transfer
85
of bacterial genes into the eukaryal nucleus have occurred after
the endosymbiosis event at the origin of mitochondria, but they
have not displaced their eukaryal functional analogues.
Remnants of the bacterial genome remain in most mitochondria
and Remnants of the bacterial membrane is still present in
all Eukarya (either as mitochondrial or hydrogenosome
membranes).
The second possibility is to imagine that the eukaryal nucleus
originated from the ICM of an akaryotic PVC bacterium after
the formation of FECA (Fig. 3B). Again, this involves a complex
and unlikely series of events. In a first step, the archaeal
symbiont should lost its membrane, whereas in a second step,
the archaeal chromosome became enclosed by a new (double)
membrane formed via invagination of the ICM. During that
process, most of the genome of the PVC bacterium should have
been lost, except for some genes that were transferred to the
newly formed nuclear genome of archaeal origin. The latter
should have included in particular the genes for bacterial lipid
biosynthesis and those for tubulin and MC proteins.
In both scenarios, the disappearance of the thaumarchaeal
membrane is especially difficult to imagine. The complete
disappearance of the membrane of the symbiont has never
been observed in actual symbiosis (for instance mitochondria
and chloroplasts have retained the two membranes of their
bacterial ancestors) and cannot be justified easily (except by
the fact that we have to explain the present situation!). The
disappearance of the archaeal membrane is especially strange
considering that Eukarya have only conserved the archaeal
version of all universal proteins working at the membrane,
such as the V-ATPase, the Sec core proteins, or else the signal
recognition ribonucleo-particle that target to the membrane
ribosomes synthesizing membrane proteins. The Eukarya also
retain archaeal specific machinery, such as the ESCRT
proteins, to work now with a bacterial membrane.
A third possibility is to imagine that the eukaryal chromosome evolved directly from the archaeal one that remained
in the thaumarchaeon (Fig. 3C). This implies that the archaeal
membrane was transformed in situ into a bacterial one. In that
case, one should imagine that the archaeal genes for lipid
biosynthesis were specifically replaced in situ (in the archaeal
chromosome) by genes for bacterial lipid biosynthesis coming
from the PVC bacterium. Again, there was no obvious selection pressure for this replacement. For that very reason, I
previously refuted in the introduction the possibility that the
thaumarchaeon was the host in the fusion. Furthermore, in that
third scenario, one has now to explain the disappearance of the
bacterial chromosome. Of course, considering the role of
viruses in our hypothesis, one can still imagine that a virulent
virus originally infecting a PVC bacterium find its way into
a descendant of FECA and specifically destroy the bacterial
chromosome (the archaeal one being protected by the archaeal
membrane or the proto-nuclear envelope or else by specific
chemical modifications). One should then assume that this
destruction only occurred after some critical bacterial genes
had already been transferred to the archaeal nucleus (i.e. genes
encoding CM proteins and proteins for the biosynthesis of
bacterial phospholipids). This again sounds as an ad hoc
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P. Forterre / Research in Microbiology 162 (2011) 77e91
assumption. Two more problems plague this third scenario,
firstly, the endoplasmic reticulum originated now after the
nucleus, as an extension of the nuclear envelope. This is in
contradiction with sound logic, the nuclear envelope being
instead clearly an extension of the endoplasmic reticulum
(Jékely, 2003, 2007; DeDuve, 2007). Secondly, the ICM of
the PVC bacterium has no more any role in the origin of the
eukaryal nucleus, which is in contradiction with the original
idea behind the PTV scenario.
In summary, the transformation of a FECA born from
a fusion event into a reasonable ancestor of modern eukarya
remains difficult to imagine in a convincing way in the PTV
scenario as in all other fusion scenarios.
Another major problem of all fusion hypotheses (rarely
discussed) is the origin of eukaryal viruses. This problem is of
course central in the PTV scenario since the descendants
of FECA are supposed to be invaded by specific lineages
of eukaryal viruses. Where did they come from? In any
fusion scenario, eukaryal viruses should have originated from
a mixture of archaeal and bacterial viruses (Koonin et al., 2006),
in the PTV scenario, eukaryal viruses should have specifically
evolved mostly from bacterial ones, since FECA originally only
contained receptors for bacterial viruses at its cell surface. The
only archaeal viruses that might have contributed were those
that were integrated as “proviruses” in the thaumarchaeal
symbiont. Some eukaryal and bacterial viruses harbour indeed
major capsid proteins (MCPs) with similar folds. This is the
case for Herpes viruses and “head and tails” bacterioviruses
whose MCPs harbour the Hong-Kong fold, or else for NCLDV,
Adenovirus and Tectiviridae (bacterioviruses) whose MCPs
harbour the double jelly roll fold (Bamford, 2003; Bamford
et al., 2007). The replicases (RNA dependent RNA polymerase)
of some eukaryal and bacterial RNA viruses or else the proteinprimed DNA polymerases encoded by different viruses infecting either Bacteria or Eukarya are also homologous between the
two domains (Filée et al., 2002,Kidmose et al., 2010). However,
despite these homologous molecular features, eukaryal viruses
in general differ dramatically of their bacterial counterparts
(and also of their archaeal ones) both in terms of virion architecture and genome content. Since viruses infecting modern
Eukarya were probably already there at the time of LECA, one
really wonders why the fusion of an archaeon and a bacterium
would have so rapidly triggered the evolution of “head and
tailed” viruses into Herpes viruses, or else the evolution of
Tectiviridaea into NCLDV?
More generally, considering the high number and great
diversity of eukaryal virus families, how can we imagine that
the PVC bacterium and the Thaumarchaeon at the origin of the
Eukarya were bona fide targets for ancestors of all these viral
families? Indeed, in the PTV scenario, one should imagine that
ancestors of all modern eukaryal viruses were able to infect
PVC bacteria (they should have used as hosts the early
descendants of FECA) or were integrated as proviruses into
the genome of the Thaumarchaeon. If this is the case, the
descendants of these viruses should be still there today
infecting modern PVC bacteria and Thaumarchaea. Of course,
one can postulate the existence of many unknown viruses
infecting PCV Bacteria and/or Thaumarchaea that resemble
NCLDV, retroviruses (not retroelements, but bona fide retroviruses with gag, pol, env genes) and all other viruses specific
for Eukarya. However, their presence has not yet been detected
in the genomes of PCV Bacteria and Thaumarchaea that are
already available. I predict that these eukaryal-like viruses will
not be found, and that the difficulty to imagine the transmutation of archaeal and bacterial viruses into eukaryotic ones
will remain a very weak point of all fusion hypotheses.
Finally, and this is the critical point, one has to explain why
they are today three and not two versions of ribosomes on our
planet. Fusion hypotheses in fact tend to roll history backward
by fundamentally denying the critical advance in our understanding of life evolution that has been brought about by the
development of molecular biology in the middle of the last
century. The discovery of three distinct deep evolutionary
lineages of organisms based on rRNAwas the key discovery that
led to formulate the three domains concept (Woese and Fox,
1977). This observation remains the major challenge to any
kind of fusion hypotheses. In the present scenario, it is indeed
hard to explain why the ribosomes of the bacterial host
completely disappeared after the fusion and why the archaeal
ribosomes of the symbiont experienced such an extreme
acceleration of their evolutionary rate in the eukaryotic lineage,
becoming sufficiently divergent from those of their archaeal
ancestor to be recognized as a different version (sensu Woese).
I have suggested previously in this paper that protein shuttle
between the genomes of ancient Eukarya and viruses infecting
them could have explained the accelerated evolution of some
proteins in Eukarya. However, this cannot be applied in the case
of ribosomal proteins since, by definition, viral genomes do not
encode ribosomes (Raoult and Forterre, 2008). The nature of the
eukaryal ribosome thus remains central to discuss the validity of
fusion hypotheses. Proponents of fusion hypotheses will argue
that the rate of evolution of ribosomal proteins in Eukarya has
been dramatically increased by the addition of new proteins and
rRNA segments during the transition from the FECA to the
LECA. However, addition of new proteins and rRNA segments
should have instead introduce more constraints (from proteineprotein and protein-RNA interactions) to the evolution of
eukaryotic ribosomal RNAs and proteins, slowing down their
rate of divergence instead to produce an acceleration.
A particular aspect of all fusion hypotheses that make me
suspicious is that they all invoke some kind of miracle that
occurred only once in life evolution (Koonin, 2007), although
the condition for its appearance have been there for the last 3
billion years (the coexistence of some Archaea and Bacteria in
the same biotope). If such fusion was possible once, it is not
clear why many eukaryal-like lineages did not emerge
continuously during the history of our planet leading to the
emergence of multiple independent (polyphyletic) eukaryal (or
eukaryal-like) groups. One cannot argue that these lineages
have been all eliminated by the descendants of LECA, because
new eukaryal-like lineages should still appeared today.
The only possibility to solve this problem is to argue that,
for some reason, the fusion was only possible in environmental
conditions that do not exist anymore on our planet. However,
P. Forterre / Research in Microbiology 162 (2011) 77e91
in that case, one should abandon any hope to find the actual
partners and, as a consequence, to understand the reasons
which selected for the fusion event itself. Again, it seems that
we want Archaea and Bacteria to associate only because we
need an explanation for our own presence in this world.
Nevertheless, a miraculous event at the origin of this association does not seem to be a good starting point from a scientific point of view.
5. Some aspects of this novel fusion scenario that are
relevant for hypotheses based on the three domains
concept
This paper can be read as an “exercice de style” to
demonstrate the failure of the best possible fusion hypothesis
to solve all problems that plague this type of evolutionary
scenario. Another objective was to consider some aspects of
the origin of Eukarya with new angles, trying to refute as
much as possible the usual critiques put forward against fusion
hypotheses in general, and, during this process, to get new
ideas that could be generally relevant for scenarios describing
ancient cellular evolution. I am indeed tempted to conclude
from this exercise that, focusing on Thaumarchaea and PVC
bacteria on one side, and viruses on the other, bring new ideas
for more realistic scenarios for the origin of Eukarya.
The discovery of previously ignored “eukaryal-like”
features in PVC bacteria and Thaumarchaea suggests that the
Last Bacterial Common Ancestor (LBCA) and the Last
Archaea Common Ancestor (LACA) were probably both more
complex than previously thought. Some of these features might
have been even present in LUCA (see below). It is thus now
easier than before to derive modern Eukarya from LUCA and/or
from the last common ancestor of Archaea and Bacteria.
The detection of eukaryal-like MC proteins in PVC bacteria
is an especially important milestone. Phylogenetic analysis of
PVC MC proteins suggests that the PVC ancestor was probably
compartmentalized, harbouring at least four genes encoding
MC proteins (Santarella et al., 2010). Also presently restricted
to a subgroup of PVC bacteria, the presence of eukaryal-like
tubulins in some members of this superphylum might be also
significant. If PVC bacteria occupy a basal position in bacterial
phylogeny, as suggested by the analysis of slowly evolving
positions in rRNA genes (Brochier and Philippe, 2002), one can
imagine that the LBCA was itself compartmentalized, already
containing ancestors of MC proteins and tubulin. Some
observations suggest indeed a tendency to reductive evolution
in Bacteria. In particular, phylogenomic analysis suggests that
the LBCA had a complete operon for genes encoding the
typical bacterial cell division and peptidoglycan synthesis
apparatus, whereas many modern bacteria (especially in the
PVC superphylum) have only retained a subset of these genes
(Pilhofer et al., 2008).
A tendency to reductive evolution is also apparent in
Archaea. The distribution of actin, FtsZ and ESCRT proteins in
this domain suggests that LACA had a machinery for cell
division and vesicle production more complex than those of
modern archaea (Makarova et al., 2010). Similarly,
87
phylogenomic analysis indicates that the ribosome of LACA
contained more ribosomal proteins than the ribosomes of
modern archaea (Lecompte et al., 2002). Recently, using
a probabilistic model that takes into account various modes of
evolution, Csurös and Miklós, (2009) also concluded that
archaeal evolution occurred mainly via streamlining of a large
ancestral genome.
Archaea and Bacteria have thus both probably experienced
reductive evolution during the divergence of their various
phyla. If this tendency was already present in the lineages
leading to the LACA and the LBCA, it might well be that
these two ancestors were themselves already less complex
than LUCA. In particular, CM proteins from PVC bacteria and
Eukarya being probably homologous, they might have been
already present in LUCA, and later on lost in Archaea and
most Bacteria. The frequent loss of these proteins that
occurred during PVC diversification supports such a scenario
(Santarella et al., 2010; Forterre and Gribaldo, 2010). The
phylogeny of true tubulins, with tubulins of PVC bacteria
branching as an outgroup of eukaryal tubulins (Jenkins et al.,
2002) can be also interpreted as supporting the presence of
tubulin in the common ancestor of Eukarya and Bacteria.
LUCA might have thus been a rather complex compartmentalized cell (Forterre and Gribaldo, 2010).
There is presently a debate about the nature of the genome
of LUCA (either RNA or DNA). At this point, one should
realize that a compartmentalized LUCA with an ancestral
cytoskeleton does not refute the hypothesis of a LUCA with an
RNA genome. Indeed, as previously argued by Poole and
Logan (2005) RNA genomes can be replicated and repaired
faithfully. Furthermore, modern RNA viruses encode proteins
as sophisticated as those encoded by DNA viruses and some of
these proteins are very efficent in the manipulation of both the
eukaryal cytoskeleton and endomembrane system to produce
viral factories (Novoa et al., 2005; Netherton et al., 2007).
If LUCA was already a sophisticated cell with some
features of Eukarya (and probably “bacterial type” lipids) the
path from LUCA to LECA and modern Eukarya becomes
much easier to imagine without the need to invoke dubious
fusion scenarios. In the framework of the three domains
concept, modern Eukarya did not evolved from Archaea (or
from Bacteria, as in Cavalier-Smith’s scenarios, see CavalierSmith, 1989, 2010) but from an ancestor that they share with
Archaea and/or Bacteria (Fig. 4). This explains why Eukarya
exhibit a mixture of archaeal and bacterial features (we don’t
need to invoke fusion to explain this observation). Eukarya
have inherited these features either from LUCA (they were
lost in Archaea or Bacteria) or from the specific common
ancestor they share with either Archaea or Bacteria (Fig. 4).
The main advantage of the three domains scenarios is to
readily explain the existence of eukaryal specific protein
families and molecular mechanisms. These eukaryal specific
features correspond either to ancient features that have been
lost in both Archaea and Bacteria or to features that have
appeared in the eukaryal lineage after its divergence from the
two others. Unfortunately, it is not easy to decide between
these alternatives. For instance, a major question mark that
88
P. Forterre / Research in Microbiology 162 (2011) 77e91
Fig. 4. Update evolutionary scenario based on the three domains concept.
Eukarya can exhibit a mixture of features that appear today as bacterial-like
(green circles) or archaeal-like (red circles) if these features are ancient traits
that were already present in LUCA and were later on selectively lost in either
Archaea or Bacteria. In addition, Eukarya can exhibit features in common with
their sister group in the universal tree of life (orange circles). In this drawing,
the root has been put in the bacterial branch as currently assumed. In the viral
hypothesis for the origin of eukaryal complexity, many specific eukaryal
features were introduced by viruses in the eukaryal lineage (violet circles).
remains open is the existence (or not) of spliceosomes in
LUCA. Phylogenomic analyses have shown that LECA
already has a full fledge spliceosome and that its genome was
probably full of introns (Collins and Penny, 2005; Koonin,
2009). The spliceosome might thus have been either already
present in LUCA or emerged between LUCA and LECA. The
spliceosome is often view as a recent features that only originated in the eukaryal lineage, however, the recent finding of
a nucleomorph (relic nuclei of algal endosymbionts of nonphotosynthetic Eukarya) that have lost all introns and genes
encoding spliceosomal components (Lane et al., 2007) testifies
that one can easily imagine that the spliceosome was present
in LUCA and later on lost in Archaea and Bacteria.
At that point, it still remains to understand why Bacteria
and Archaea evolved according to a reduction scenario,
whereas Eukarya evolved according to a scenario whose
hallmark is sophistication (at least from FECA to LUCA, since
many eukaryal lineages have experienced reductive evolution
after LECA). I suggested 15 years ago that Bacteria and
Archaea evolved toward simplification because they experienced passage through a thermophilic period (Forterre, 1995).
In that “thermoreduction hypothesis”, their “prokaryotic”
characters (small size, high macromolecular turnover,
coupling of traduction and translation) were selected in
response to adaptation to high temperatures (see also Poole
et al., 1999; Glansdorff et al., 2008). A major problem for
life at high temperature is indeed the chemical thermodegradation of informational macromolecules, especially mRNA
and proteins. Thermoreduction might have been stronger in
Archaea, with, in particular, the invention of “archaeal lipids”
that, together with those of reverse gyrase (Forterre, 2002),
allowed some of them to conquer biotopes above 90 C.
The thermoreduction hypothesis has recently received
a strong experimental support from the work of Boussau and
co-workers who concluded from in silico analyses of rRNA
and universal proteins evolution that LUCA was probably
a mesophile, but that the LACA and LBCA were both thermophiles (Boussau et al., 2008). In that case, Eukarya might
be the descendants of those ancient cells that failed to adapt
to high temperature. Most of them were possibly wiped out
by the descendants of the LACA and LBCA that adapted later
on to low temperature, except those with large size that
succeeded to make a living by eating mesophilic archaea and
bacteria.
Another possibility (non-exclusive) is that viruses play
a role in the orientation of each domain toward different
evolutionary pathways. In that case, we can reintroduce here
our proposals on the role of viruses in the PTV scenario for
the origin of Eukarya, but this time in the framework of the
three domains concept (Fig. 4). One can still assume that
massive invasions of ancient Eukarya by specific viral families bring indeed new proteins in the eukaryal lineage and
induced an arms race that favoured their evolution toward
increasing complexity. The ideas that many eukaryal specific
proteins originated first in NCLDV or other viral lineage
unique to Eukarya and that retroviruses played a critical role
in shaping the eukaryal chromosomes and/or in the origin of
meiotic sex can be easily recycled in scenarios based on the
three domains concept. Furthermore, whereas in fusion
hypotheses eukaryal viruses are supposed to have evolved
from bacterial or archaeal viruses, something difficult to
accept, one can suggest now that specific eukaryal viruses,
such as retroviruses and NCLDV, originated from ancient
viruses that were already thriving at the time of LUCA
(Prangishvili et al., 2006a).
As previously mentioned, a striking conclusion from studies
of viral diversity is that members of different domains are
infected by different viral families (for review, see Prangishvili
et al., 2006a). In particular, at least in the state of our present
knowledge, Eukarya are infected by a much greater diversity
of viral families than Bacteria and Archaea. For instance,
RNA viruses are very abundant and diverse in Eukarya (singlestranded, double stranded, segmented, non segmented) whereas
they are quite rare in Bacteria and presently unknown in
Archaea. In the framework of the thermoreduction hypotheses,
it is possible that RNA viruses (relics of the RNA-protein world,
see Forterre, 2005) were unable, with few exceptions, to
follow their hosts in hot environments, considering the high
instability of RNA at high temperature (Forterre, 1995). In
particular, retroviruses and most retroelements, that are so
abundant in eukaryal genomes, might have been eliminated
during the period of reductive evolution that led from LUCA
to LBCA and LACA.
P. Forterre / Research in Microbiology 162 (2011) 77e91
I suggest here that the type of viruses that were “selected”
by a given lineage at the onset of the formation of the three
domains of life (Prangishvili et al., 2006a) was determinant in
the mode of evolution of this domain. Hence, the type
of viruses present in ancient Eukarya was possibly a major
factor in the evolution of this domain toward more and more
sophistication. In particular, the loss of retroviruses in Archaea
and Bacteria and their conservation in Eukarya might have
been a major factor (possibly in addition to the thermoreduction story) that pushed the evolution of Archaea and
Bacteria toward reduction and those of Eukarya toward complexification. The absence of retroviruses in yeast could
explain, for instance, why these microbial Eukarya, which
have evolved by reduction from multicellular fungi, seem now
“blocked” in a kind of “prokaryotic-like situation” (small size,
small and compact genomes, few introns, unicellular).
All models for the origin of Eukarya have generally ignored
the possible role of viruses in that process, except for the origin
of the nucleus (Takemura, 2001; Bell, 2001). However,
comparative genomics and common sense tell us that, in order to
understand the history of life on our planet, one has to study not
only the evolutionary relationships between the three cellular
domains, but also those between the three viral “domains”, and,
above all, the consequences of their co-evolution. For that
reason, I am really convinced that the viral hypothesis for the
origin of complexity in Eukarya is worth further exploration.
6. Perspective
Hypotheses on the origin of Eukarya will be always based
on our knowledge of modern biodiversity and molecular
mechanisms. The discovery of novel eukaryal features in
groups of Archaea and Bacteria only recently recognized and
extensively studied should encourage us to continue actively
the exploration of the microbial world. The recent discovery of
giant multicellular species in Thaumarchaea illustrate well the
possibility of further surprising discoveries in these phyla
(Muller et al., 2010). The discovery of mimivirus was another
breakthrough that helped to remind biologists of the importance to consider viruses in evolutionary scenarios (Raoult
et al., 2004; Claverie and Abergel., 2010, Forterre, 2010a).
In-depth exploration of the cellular and viral biodiversity,
especially in those microbial phyla for which there is presently
no cultivable representatives, should be now on the top priority
list of biologists looking for exciting scientific adventures in
the XXI century (and of course on the top priority list of
funding agencies and Heads of Institutes). We don’t know if
we will have one day the last word on the origin of Eukarya,
but, if coupled to further exploration of terrestrial life, this
quest will certainly not be vain. In any case, one can only
wonder how many surprises remain to be unveiled in the vast
microbial world.
Acknowledgments
I am grateful to the two reviewers for especially important
remarks and suggestions.
89
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