Horizontal gene transfer in eukaryotic algal evolution

Abstract

Although seemingly innocuous, the power of something as simple as a drop of water, given a few millions of years over which to act continually, is immediately and awe-inspiringly obvious to someone peering over the edge of the Grand Canyon. Acting over a similarly immense time period but on vastly different scales, this massive effect from a weak, but constant, force embodies how natural selection has been able to direct the evolution of once primitive, mal-adapted biological structures to the remarkable and almost inconceivably diverse molecular machines found within extant organisms. This idea of natural selection as a slow-and-steady workhorse was central to Charles Darwin's evolutionary synthesis, as epitomized in his oft-repeated precis ``Natura non facit saltum,'' Nature does not make leaps. Darwin would not live to see the discovery of genes as the vessel of inheritance and random mutation as the propagator of change, although these breakthroughs would serve to reinforce his prescient ideas.

Full text

Horizontal gene transfer in eukaryotic algal evolution
Jason Raymond and Robert E. Blankenship*
Department of Chemistry and Biochemistry, Arizona State University, Tempe, AZ 85287-1604
Although seemingly innocuous,
the power of something as sim-
ple as a drop of water, given a
few millions of years over
which to act continually, is immediately
and awe-inspiringly obvious to someone
peering over the edge of the Grand
Canyon. Acting over a similarly im-
mense time period but on vastly differ-
ent scales, this massive effect from a
weak, but constant, force embodies how
natural selection has been able to direct
the evolution of once primitive, mal-
adapted biological structures to the re-
markable and almost inconceivably di-
verse molecular machines found within
extant organisms. This idea of natural
selection as a slow-and-steady workhorse
was central to Charles Darwin’s evolu-
tionary synthesis, as epitomized in his
oft-repeated pre´cis ‘‘Natura non facit
saltum,’’ Nature does not make leaps.
Darwin would not live to see the discov-
ery of genes as the vessel of inheritance
and random mutation as the propagator
of change, although these breakthroughs
would serve to reinforce his prescient
ideas.
To the contrary, the discovery of hor-
izontal gene transfer (HGT) as a signifi-
cant evolutionary driver may require an
addendum to the Darwinian synthesis.
A growing body of evidence indicates
that many organisms, particularly pro-
karyotes, can and do make evolutionary
leaps by sharing genes with one another,
thereby opening a back door to an ad-
aptation or ability that was already fine-
tuned within another organism. Once
thought to be an explanation of last re-
sort when the data were not robust
enough to give unambiguous results,
with the recent availability of a wealth
of whole-genome data, HGT has not
only become respectable but has
emerged as a central force in the evolu-
tion of many different prokaryotes
(1–3). Of course, this idea came as
no major surprise to many bacterial
geneticists, who for decades have been
selecting prokaryotes for their ability to
take up and express exogenous genes (as
Oswald Avery did some 60 years ago,
demonstrating that DNA was the carrier
of genetic information) (4).
The impact of HGT on eukaryote ge-
nomes has not been so clear-cut (3).
The species concept of genetically segre-
gated germ lines has been tied to eu-
karyote taxonomy since its inception,
and the barriers against HGT in bacte-
ria are magnified in eukaryotes by fur-
ther complexities in transcription and
translation, such as the need for correct
splicing of RNA transcripts replete with
introns. It can also be argued that sexual
reproduction affords many eukaryotes
the same advantage gained through
HGT in bacteria. In this issue of PNAS,
Archibald et al. (5) leap beyond the
case-by-case examples that typify eu-
karyotic HGT and demonstrate that
HGT has played a significant role in the
evolution of a eukaryotic alga. In a col-
lective analysis of 78 plastid-targeted
proteins from this alga, they show that,
even by conservative measures, ⬇21%
of these genes have likely been acquired
by HGT. Their result stands to signifi-
cantly expand the number of established
cases of so-called transdomain HGT oc-
curring between prokaryotes and eu-
karyotes and bolsters some novel ideas
on evolutionary mechanisms in phago-
cytic eukaryotes (6).
The subject, Bigelowiella natans,isa
member of a class of algae known as
chlorarachniophytes that, in and of it-
self, is quite an evolutionary enigma. All
plastid-containing eukaryotes acquired
the ability to do photosynthesis when,
perhaps ⬇2 billion years ago, a primitive
eukaryote engulfed a photosynthetic
cyanobacterium. This so-called primary
endosymbiotic event gave early eu-
karyotes an extremely powerful meta-
bolic ability that previously was manifest
only among photosynthetic bacteria, and
also constituted, along with the enslave-
ment of a proteobacterium that would
become the mitochondrion, a massive
horizontal transfer of genes into a prim-
itive eukaryote. The modern progeny of
this primitive photosynthetic eukaryote
point to a single primary endosymbiotic
event, although some evidence argues
otherwise (e.g., refs. 7 and 8). It is also
certainly feasible that endosymbiosis
occurred multiple times, but many or-
ganisms were wiped out in the bottle-
neck of subsequent global catastrophes
(e.g., global glaciations, refs. 9–11).
Not to be left behind, some eu-
karyotes acquired photosynthesis
through the same mechanism, although
not by engulfing a cyanobacterium but
rather a eukaryotic alga (10, 12).
Termed secondary endosymbiosis, this
process is believed to have given rise to
multiple independent groups of photo-
synthetic organisms, all of which bear
the hallmark of plastids with three or
more bounding membranes (13). That
secondary endosymbiosis has occurred
multiple times is also made clear in that
it has occurred in various lineages after
the radiation of the three flavors of pri-
mary algae (red, green, and glaucocysto-
phyte), leading to secondary algae with
quite varying plastid phenotypes. In pri-
mary and secondary endosymbiosis
there has been a massive loss of genes
from the endosymbiont genome, many
of which have been transferred into the
host genome, with each host-encoded
plasmid-targeted gene now carrying a
transit peptide sequence that directs it
back to the plastid (secondary endosym-
biont-directed proteins also carry an
additional signal sequence to get them
through the vestigial membrane system
of the original plastid host) (14). Two
groups of algae that evolved through
secondary endosymbiosis, the crypto-
mondads and chlorarachniophytes (in-
cluding B. natans) are particularly inter-
esting because they still contain a relict
nucleus called a nucleomorph, dramati-
cally reduced in size, from the originally
engulfed algae (10).
Additional transfer of genes has un-
doubtedly occurred from the relict nu-
cleus into the host genome (15), al-
though despite these complex transfer
events these genes in the host genome
should have a phylogenetic signal consis-
tent with the engulfed algae and thus
should be grouped more broadly with
cyanobacteria. Genes present in the host
before endosymbiosis should cluster
with other eukar yotic genes and thereby
can be used to classify the original host.
For most nuclear-encoded algal genes
these stratifications are indeed observed,
and in B. natans a variety of evidence
clearly indicates a green algal endosym-
biont origin. However, as Archibald
et al. (5) show, many of the plastid-
targeted genes from B. natans clearly
diverge from this expectation. These
horizontally transferred genes span a
varied swath of functions, including
chlorophyll biosynthesis, carbon fixation,
and ribosome structure, and cluster with
a similarly broad range of taxa other
than green algae. Several of their trees,
which, importantly, encompass much of
the available taxonomic sampling for
each sequence, are particularly robust
based on bootstrap values and con-
served sequence motifs, providing strong
See companion paper on page 7678.
*To whom correspondence should be addressed. E-mail:
blankenship@asu.edu.
www.pnas.org兾cgi兾doi兾10.1073兾pnas.1533212100 PNAS
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no. 13
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COMMENTARY

support for HGT having played a signif-
icant role in the evolution of this
organism.
Because all of the genes studied by
Archibald et al. are encoded within the
nucleus of B. natans but operate within
the plastid, the signal and transit pep-
tides are absolutely necessary. One can
surmise that this necessity would
strongly favor HGT in and among algae,
where nuclear-encoded genes targeted
to plastids must navigate a similar maze
of endomembranes through the direc-
tion of transit peptides, and additional
signal peptides in the case of other sec-
ondary algae. Indeed, a majority of
genes studied by the authors support
this expectation, favoring phylogenies
consistent with HGT from streptophytes
or red algae into the B. natans genome.
Most intriguingly, two of the genes from
their analysis indicate HGT from differ-
ent bacteria, significant not only as an
example of prokaryote-to-eukaryote
gene transfer but also because these ac-
quired genes initially would have not
had the proper leader sequence for
import into the plastid. Whether the
appropriate targeting sequence was in-
corporated de novo through gene con-
version or some other mechanism of
homologous or orthologous replacement
is not clear, but this remarkable finding
certainly invokes new ideas on how
genes are assimilated into a genome.
Microbiologists have long known
about phenotypes that favor promiscu-
ous plasmid sharing among bacteria,
responsible for the epidemic spread of
antibiotic resistance. Although no plas-
mid analog exists in eukar yotes,
Archibald et al. suggest that HGT in B.
natans may occur in the same way it has
for the many endosymbiotic events that
have happened over the past 2 billion
years, by engulfing other organisms (Fig.
1). Compared with the green alga
Chlamydomonas reinhardtii, which is
photoautotrophic and in which no paral-
lel evidence of HGT is found, B. natans
is mixotrophic, meaning that it can live
phagocytotically and photosynthetically.
Models have been proposed whereby
small snippets of DNA from an en-
gulfed microbe are able to escape diges-
tion, e.g., from protist lysozomes, and
migrate to and subsequently be incorpo-
rated into the host genome (6, 16). One
can imagine the series of fleetingly small
probability events proceeding from en-
gulfment to incorporation of a strand of
foreign DNA into the genome to a new
gene overcoming genetic drift to be-
come fixed in the population. In a cer-
tain sense, this is the same cumulative
effect as random mutations in single
genes or dripping water, but it now op-
erates on a different level, an entire
gene. However, the essential point is
that those probabilities are nonzero and
over time have made a significant con-
tribution to the genome of this organism
(6). The real boon of Archibald et al.’s
hypothesis, perhaps best synopsized by
Ford Doolittle’s epigram ‘‘you are what
you eat’’ (16), is that it is eminently test-
able as more eukaryotic genome data
become available. It is already apparent
that the magnitude of HGT varies dra-
matically in different lineages of algae,
with the proposed explanation of the
phagocytotic lifestyle as a likely but not
proven explanation for the observed
mosaic pattern. Whether this is a more
general mechanism for HGT in a wider
range of eukaryotes, including nonalgal
taxa, is not yet apparent.
So although Nature herself may not
make leaps, it now seems clear that
many organisms, eukaryotes and pro-
karyotes, are certainly able to mimic
evolutionary jumps through HGT.
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Fig. 1. Stepwise conceptual image of one mechanism of HGT that, as proposed by Archibald et al. (5),
might operate in the chlorarachniophyte alga B. natans. Red arrows show phagocytosis and subsequent
digestion of a bacterium or protist, from which foreign DNA has survived digestion and become
incorporated into the algal nucleus (flow of HGT-acquired genetic information indicated with blue
arrows). Although the genes studied herein by Archibald et al. are directed for function to the plastid, the
significant number of horizontally transferred genes they found may only be the tip of the iceberg in
phagocytic protists such as B. natans.
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www.pnas.org兾cgi兾doi兾10.1073兾pnas.1533212100 Raymond and Blankenship