See discussions, stats, and author profiles for this publication at: https://www.researchgate.

net/publication/8987066

Mitochondrial genomes: anything goes. Trends Genet 19:709-716

Article  in  Trends in Genetics · January 2004

DOI: 10.1016/j.tig.2003.10.012 · Source: PubMed

CITATIONS READS
441 424

3 authors, including:

Gertraud Burger
Université de Montréal
274 PUBLICATIONS   12,639 CITATIONS   

SEE PROFILE

Some of the authors of this publication are also working on these related projects:

Perfection of eccentricity: Mitochondrial genomes of diplonemids View project

Mitochondrial Genome Structure View project

All content following this page was uploaded by Gertraud Burger on 30 June 2018.

The user has requested enhancement of the downloaded file.

 Review
Mitochondrial genomes: anything goes
Gertraud Burger1, Michael W. Gray2 and B. Franz Lang1
1
Canadian Institute for Advanced Research, Programme in Evolutionary Biology, Départment de Biochimie, Université de Montréal,
2900 Boulevard Edouard-Montpetit, Montréal, Québec, Canada H3T 1J4
2
Canadian Institute for Advanced Research, Programme in Evolutionary Biology, Department of Biochemistry and Molecular
Biology, Dalhousie University, Halifax, Nova Scotia, Canada B3H 1X5
Mitochondria have their own genetic system – a vesti-
gial genome originating from an endosymbiotic
a-proteobacterial ancestor. The genetic function of
mitochondrial DNA (mtDNA) is well-conserved, being
involved in a maximum of five mitochondrial processes:
invariantly in respiration and/or oxidative phosphoryl-
ation and translation, and also in transcription, RNA
maturation and protein import. Recent data from mito-
chondrial genomics research demonstrate that this gen-
etic conservatism contrasts with an often-perplexing
diversity in structure and gene expression mechanisms.
In addition to outlining the extraordinary diversity of
mtDNA, this review highlights the divergent trends in
mitochondrial genome evolution in the various eukary-
otic lineages, and examines the relationship between
mitochondrial and nuclear genome evolution in a given
organism.
Mitochondria, the morphologically distinctive double-
membrane organelles of most cells, use electron transport
coupled with oxidative phosphorylation to generate ATP
[1]. Although mitochondria have a central role in energy
transduction (the powerhouse of eukaryotic cells), these
organelles participate in several other important func-
tions, including ion homeostasis, intermediary metab-
olism and apoptosis. However, in only a few instances do
these pathways have components encoded by mitochon-
drial DNA (mtDNA) (Figure 1); rather, most mitochondrial
proteins are specified by nuclear genes, synthesized in the
cytosol and imported into the organelle [2].
Mitochondria are ubiquitous throughout the eukaryotic
domain: with very few exceptions, eukaryotes cannot exist
without them. Eukaryotic lineages that lack both func-
tional mitochondria and a mitochondrial genome do exist,
but several of these ‘amitochondriate’ eukaryotes contain
DERIVED (see Glossary) mitochondria (e.g. hydrogeno-
somes) that generate ATP anaerobically [3]. Other
amitochondriate species contain what appear to be
remnant mitochondrial structures (of currently undefined
function), and/or express nucleus-encoded proteins typi-
cally targeted to, and functioning in, mitochondria [4].
These observations argue that the last common ancestor of
extant eukaryotes was a mitochondria-containing organ-
ism, implying that PRIMITIVE amitochondriate eukaryotes
(eukaryotes whose ancestors never had mitochondria)
might not exist.
Corresponding author: Gertraud Burger (gertraud.burger@umontreal.ca).
http://tigs.trends.com 0168-9525/$ - see front matter q 2003 Elsevier Ltd. All rights reserved. doi:10.1016/j.tig.2003.10.012
TRENDS in Genetics Vol.19 No.12 December 2003 709
The limited genetic function of mtDNA is well-con-
served across eukaryotes: mtDNA encodes a small number
of proteins whose mRNAs are translated by a distinctive
mitochondrial protein-synthesizing system, some of whose
components are always (rRNAs), usually (tRNAs) or
occasionally (ribosomal proteins) specified by the mito-
chondrial genome. In marked contrast to this genetic
conservatism (as this review will emphasize), the mito-
chondrial genome is characterized by an extreme and often
bewildering diversity in structure and gene expression
mechanisms [5,6]. This dichotomy – genetic conservatism
versus structural flamboyance – has long obscured the
origin of mitochondria and raised the question of whether
this organelle arose more than once during eukaryotic cell
evolution. The weight of evidence now firmly argues that
the mitochondrion had a single origin, arising from a
eubacterial symbiont whose closest contemporary rela-
tives are found within the a-proteobacteria.
This review summarizes recent data from mitochon-
drial genomics research. It aims to highlight the extra-
ordinary structural diversity shown by mtDNA across the
eukaryotic domain (Figure 2), and the exceptionally
divergent trends in mitochondrial genome evolution in
various eukaryotic lineages. Chloroplasts, a second type of
DNA-containing organelle in algae and plants, will not be
discussed. It is worth noting that chloroplasts, which are
thought to be the product of a more recent endosymbiotic
event than mitochondria, have genomes that show
significantly less structural diversity than mitochondrial
genomes.
The many faces of mitochondrial genome architecture
Given that the mitochondrial genome of several animals
exists in the form of super-coiled DNA [7], and that mtDNA
in most eukaryotes maps as a circle in restriction analysis
and sequence assembly, it was assumed, until recently,
that mtDNAs are generally circular molecules. However,
there is now strong evidence that many, if not most,
Glossary
Derived: taxa or characters that have evolved far away from the primitive state.
In molecular phylogeny, long tree-branches indicate highly derived taxa.
Homologous: similar in sequence owing to common evolutionary origin.
Primitive: original or ancient state.
Monophyly: unique origin in a single ancestral species (a monophyletic
grouping includes the ancestor and all of its descendants).
Basal (early diverging): emerging early in the evolution of a clade (i.e. a deep
branch in a phylogenetic tree).
710 Review TRENDS in Genetics Vol.19 No.12 December 2003

Lipid metabolism Heme synthesis Proteases

Nucleotide metabolism Fe-S synthesis Chaperones
Amino acid metabolism Ubiquinone synthesis Signalling pathways
Carbohydrate metabolism Co-factor synthesis DNA repair, replication, etc.

RNA Heme lyase RNAse P

polymerase

EF-Tu Ribosome

TIM Oxa1
translocases translocase

Sec Tat
translocase translocase

Complex I Complex V
Complex II Complex IV
Complex III
Cytochrome c
TRENDS in Genetics

Figure 1. Biological processes in mitochondria. The majority of mitochondrial functions is shown; only the mitochondrial matrix and inner membrane are depicted.
(For more details on mitochondrial metabolism and transport see [79].) Most mitochondrial components are encoded by the nucleus (blue); those highlighted in pink are
encoded by mtDNA in some eukaryotes but by the nuclear genome in other eukaryotes, whereas a small portion is invariably specified by mtDNA (orange). Processes that
have exclusively nucleus-encoded constituents are listed. Complexes I– V are involved in electron-transport and oxidative phosphorylation. TIM translocases are involved
in protein import and insertion into the inner membrane. Tat, Sec and Oxa1 translocases are involved in protein export from matrix and insertion into the inner membrane
(for a review see [2]). RNase P is a ribozyme that processes the 50 -end of tRNAs (for recent results on fungal RNase P RNAs see [42]). It should be noted that the majority of
eukaryotes has a multi-subunit, rotenone-sensitive NADH dehydrogenase as depicted, with nad1, nad4 and nad5 invariantly encoded by mtDNA. However, in a few organ-
isms (yeast, Schizosaccharomyces and Plasmodium), this complex is replaced by a nucleus-encoded, single-polypeptide enzyme [80]. Abbreviation: EF-Tu, elongation
factor Tu.

Holozoa
Amoebidium
Monosiga Saccharomyces
Metridium
Fugu Rhizopus Fungi
Homo Allomyces
Arabidopsis Harpochytrium
Green algae Marchantia Dictyostelium
and plants Pedinomonas
Amoebozoa
Acanthamoeba
Scenedesmus Plasmodium Apicomplexa
Chlamydomonas Tetrahymena Ciliates
Cyanidioschyzon Paramecium
Red algae
Porphyra Cafeteria
Cryptomonas Ochromonas Stramenopiles
Cryptophytes Phytophthora
Rhodomonas
Reclinomonas Trypanosoma
Jakobids Euglena
Euglenozoa
Jakoba

TRENDS in Genetics

Figure 2. Eukaryotic phylogeny. Schematic phylogenetic tree depicting relationships among eukaryotes based on evidence from both statistically well-supported molecular
phylogenies with different datasets [81], and from conjecture. Only the better-known, major organismal groups are shown, with taxa belonging to the same clade sharing
the same color shade. Broken lines indicate uncertain branching orders and tree topologies. For example, the Monophyly of apicomplexa plus ciliates and stramenopiles is
not well supported. Euglenozoa and jakobids have been suggested to be the most Basal (Early Diverging) lineages; however, the relative branching order at the base of the
tree (in excess of 1 billion years) remains largely unresolved.

http://tigs.trends.com
 Review TRENDS in Genetics Vol.19 No.12 December 2003
circular-mapping mtDNAs consist primarily of linear,
multimeric head-to-tail concatamers [8,9]. Linear map-
ping (i.e. linear monomeric) mtDNA molecules have
also been detected in a few unrelated organisms, such
as ciliates, apicomplexa (Plasmodium and relatives),
fungi, chlorophycean green algae (Chlamydomonas and
relatives), and several cnidarian animals. Notably,
these linear molecules contain various specialized end-
structures, such as covalently closed single-stranded
DNA termini and terminally attached proteins; in
addition, they tend to have telomere-like repeats of
varying length [10].
Mitochondrial genomes typically consist of a single
‘chromosome’. Cases of mitochondrial genomes comprising
multiple, circular-mapping molecules have been reported
in Spizellomyecs punctatus [11], a chytridiomycete fungus,
and in the animals Globodera [12] and Dicyema [13]. A
most unusual and, indeed, unprecedented situation has
been identified in a single-celled protistan relative of
animals, Amoebidium parasiticum [14], whose mtDNA
comprises a complex set of several hundred distinct types
of linear molecules [15] (Figure 3). These molecules carry
at their termini a common pattern of short terminal
repeats, consisting invariably of the same ,40-bp term-
inal sequence motif in inverted orientation, followed
internally by an , 100-bp repeat at one end, and a
different , 65-bp repeat at the other end. The large
number of distinct ‘chromosomes’ and the apparent
absence of centromeric structures characteristic of nuclear
chromosomes raise questions about the mechanism of
concerted replication and equal segregation of mtDNA
during mitochondrial division and subsequent cell
division in A. parasiticum. A similar dilemma exists in
kinetoplastid protists (e.g. Trypanosoma), whose mito-
chondria contain up to a few dozen gene-coding maxicircles
and up to several thousand minicircles that specify the
guide RNAs involved in editing of mitochondrial
mRNAs [16].
Tetrahymena mtDNA
Human mtDNA
10 kbp
Spizellomyces mtDNA Amoebidium mtDNA
TRENDS in Genetics
Figure 3. Mitochondrial genome architectures. DNA molecules are mostly circular-
supercoiled in humans, and linear-monomeric in the ciliate Tetrahymena pyrifor-
mis (reviewed in [10]). In the fungus Spizellomyces punctatus, mtDNA consists of
three types of circular-mapping molecules [6], whereas several hundred types of
linear-monomeric molecules comprise the mitochondrial genome in the ichthyos-
porean protist Amoebidium parasiticum [15]. Triangles represent terminal repeat
motifs of 32 bp in T. pyriformis mtDNA and 40 bp in A. parasiticum mtDNA. Filled
circles and diamonds represent sub-terminal repeats, which are ,100-bp and
, 65-bp long, respectively, in A. parasiticum mtDNA.
http://tigs.trends.com
711
Size versus content
The size of mtDNA in most eukaryotic phyla ranges from
15 – 60 kbp; however, there are some notable exceptions.
Among those organisms whose mtDNA has been comple-
tely sequenced, the two extreme outliers are the apicom-
plexan protists Plasmodium sp. (obligate parasites
causing malaria), which have a minuscule mitochondrial
genome of 6 kbp [17], and rice (Oryza sativa), whose
mtDNA (at 490 kbp [18]) is , 80 times larger than that of
Plasmodium. The mapped, but as-yet unsequenced
mtDNAs of cucurbit plants could be even larger than
2000 kbp [19].
The coding capacity of mtDNA ranges from nearly
100 genes in jakobid flagellates [20] to only five in
Plasmodium, with the average across eukaryotes being
40 – 50 genes. In contrast to their up-to-20-fold differ-
ence in number, mitochondrial genes are involved, in at
most, five basic processes: invariantly in respiration
and/or oxidative phosphorylation and translation, and
occasionally also in transcription, RNA maturation and
protein import (Figure 4).
Somewhat counter-intuitively, there is no correlation
between mtDNA size and gene content (Figure 5). In fact,
the differences in size of mitochondrial genomes are mostly
caused by marked variations in the length and organiz-
ation of intergenic regions, which, in some cases, consist of
extensive tandem-repeat arrays [21,22] or stem-loop
motifs [23]. Some of the repeat elements have been
proposed to be mobile and ‘selfish’ (e.g. [24]); others, to
play a role in recombination [25]. Another determinant of
mtDNA size is gene structure: a given gene product can be
encoded by stretches of DNA of variable length owing to
the presence of introns (group I and II) of various size
(0.15 –4.00 kbp) and number (0 to .30). In the filamentous
fungus Podospora anserina, introns account for up to 75%
of the overall size of mtDNA [26].
Unusual mitochondrial gene structures
In addition to genes split by introns, mitochondria show
other, more unusual gene organizations, one of which is
gene fusion. In two amoebozoan protists, Acanthamoeba
castellanii [27] and Dictyostelium discoideum [28], the
genes coding for subunits 1 and 2 of cytochrome oxidase
(cox1 and cox2, respectively) are immediately adjacent to
one another and in the same transcriptional orientation in
the mtDNA, forming a single contiguous ORF (Figure 6a).
The existence of a bi-cistronic mRNA has been shown in
Acanthamoeba, but it is not clear at this time whether a
fusion protein is initially synthesized as well [29].
Chimeric genes, often containing recognizable pieces of a
known gene (or genes) within an otherwise unassigned
ORF, have been associated with cytoplasmic male sterility
in plants, a phenotype that is probably attributable to
alterations in electron-transport-coupled oxidative phos-
phorylation in pollen-producing cells [30]. In other cases
(e.g. [31,32]), the functional consequences (if any) of
reported gene fusions are unknown.
Genes that exist in pieces, which were first described for
the mitochondrial rRNA genes in the ciliate protozoon
Tetrahymena pyriformis [33,34] and the green alga
Chlamydomonas reinhardtii [35], represent another type
712 Review TRENDS in Genetics Vol.19 No.12 December 2003

100 400

90
350
80

Mitochondrial genome size (kbp)

70
200
Number of genes

60

50 150

40
100
30

20 50

10
0
0 is m tia ba es as as ria um mo as um
ps diu n ko yc n n te di n i
as as tia ria on sis iga mo um nas ces ium ido ebi rcha Ja om omo omo afe etri Ho mo od
on on han fete hyz dop os Ho ridi o y od ab o a r n od C M o
yd las
m
om dom arc Ca osc abi on et nom rom sm Ar Am M ha cli
n cc Re Rh am P
cli ho M id
i
Ar
M M di ha Pla a l
Re R an Pe acc
S Ch
Cy S Key:

Key: Authentic mitochondrial genes

Respiration and oxidative phosphorylation
Ribosomal RNAs Protein import and maturation
Transfer RNAs RNA maturation
Ribosomal proteins and EF-Tu Transcription
TRENDS in Genetics
Figure 4. Mitochondrial gene classes and their representation across eukaryotes.
Genes included in the functional classes are as follows. Respiration and oxidative
phosphorylation: atp1, atp3, atp4, atp6, atp8, atp9; cob; cox1–3; nad1 –4, nad4L,
nad6 –11, sdh2 –4. rRNAs: rnl, rns, rrn5. tRNAs: trnA–Y (among others). Ribosomal
proteins and EF-Tu: rps1– 4, rps7, rps8, rps10– 14, rps19; rpl1, rpl5, rpl6, rpl10,
rpl11, rpl14, rpl16, rpl18– 20, rpl27, rpl31, rpl32, rpl34, rpl36; tufA. RNA maturation:
rnpB. Protein import and maturation: secY, tatC, yejR (ccmF), yejU (ccmC), yejV
(ccmB), yejW (ccmA); cox11. Transcription: rpoA-D. Species names are: Reclino-
monas americana (jakobid flagellate); Rhodomonas salina (cryptophyte alga);
Marchantia polymorpha (liverwort, bryophyte); Cafeteria roenbergensis (strame-
nopile flagellate); Cyanidioschyzon merolae (red alga); Arabidopsis thaliana (flow-
ering plant, angiosperm); Monosiga brevicollis (choanozoan flagellate); Homo
sapiens (vertebrate animal); Metridium senile (cnidarian animal); Pedinomonas
minor (green alga, chlorophyte); Saccharomyces cerevisiae (ascomycete fungus);
and Plasmodium falciparum (apicomplexan protist). Data taken from the Organelle
Genome Database GOBASE (http://megasun.bch.umontreal.ca/gobase/).
Introns, ORFs, plasmid-derived genes
Non-coding regions
TRENDS in Genetics
Figure 5. Mitochondrial genome size and coding content across eukaryotes.
Length of coding regions of authentic mitochondrial genes (purple), introns, intro-
nic ORFs, phage-like reverse transcriptases and DNA polymerases (blue), and
intergenic regions (green). Species as in Figure 4, plus Amoebidium parasiticum
(ichthyosporean protist); Jakoba libera (jakobid flagellate); and Chlamydomonas
reinhardtii (green alga, chlorophyte).
translated into two separate polypeptides (Figure 6b);
however, post-translational protein fusion can not be ruled
out at present [37].
A stunning variation on this theme is illustrated in two
recent reports. In the green alga Scenedesmus obliquus,
the mitochondrial cytochrome oxidase subunit 2 is
apparently encoded by two gene pieces; that specifying
the N-terminal portion is located on mtDNA [38], whereas

(a) (b) (c)

of unorthodox gene structure in mitochondria. In some cox2-b
instances, these genes are broken up into as many as ncDNA
eight modules that are scrambled in the genome,
cox1 cox2 nad1-a cox2-a
interspersed with other genes and located on both mtDNA
strands of the mtDNA [35]. Separate transcription of nad1-b

these subgenic modules yields discrete pieces of rRNA
that are held together via base pairing of complemen-
tary sequence stretches. In P. falciparum, the number
of mtDNA-encoded rRNA pieces is even higher than in
Chlamydomonas [36].
The genes of a few mtDNA-encoded proteins also
display a ‘genes-in-pieces’ type of arrangement. In two
ciliate species, Tetrahymena pyriformis and Paramecium
aurelia, the nad1 gene is split into two fragments that are
transcribed into two separate mRNAs, which are probably
Acanthamoeba Tetrahymena Scenedesmus
Dictyostelium Paramecium
TRENDS in Genetics
Figure 6. Unusual structures of mitochondrial genes. (a) cox1 and cox2 genes spe-
cifying cytochrome oxidase subunits 1 and 2. (b) nad1 encodes NADH dehydro-
genase subunit 1. Mitochondrial DNA (mtDNA) and nuclear DNA (ncDNA) are
shown in blue and purple, respectively. Suffixes -a and -b indicate the gene pieces
encoding the N- or C-terminal portion of a given protein, respectively. (c) cox2
gene, specifying cytochrome oxidase subunit 2. Species are: Acanthamoeba
castellanii; Dictyostelium discoideum (amoebozoa); Tetrahymena pyriformis; Para-
mecium aurelia (ciliates); and Scenedesmus obliquus (chlorophycean green alga).

http://tigs.trends.com
 Review TRENDS in Genetics Vol.19 No.12 December 2003
the C-terminal portion is believed to be nucleus-encoded
[39] (Figure 6c). In the green algae Chlamydomonas and
Polytomella, the corresponding two protein portions are
encoded by separate nuclear genes, which are synthesized
in the cytoplasm and imported into the mitochondrion [39].
A second recent example of a split gene with one-half in
mtDNA and the other half in nuclear DNA is rpl2 in
flowering plants [40].
Finally, gene simplification is another eccentric
phenomenon seen in mitochondrial genomes. Examples
include the truncated tRNAs (lacking one, or more, helical
arms) found in the mitochondria of several animal lineages
[7], the structurally streamlined mitochondrial rRNAs in
most animals [7] and some protists [41], and mitochondrial
RNase P RNAs in fungi [42]. In certain organisms
(e.g. in the fungus Harpochytrium [43]), a similar
trend is observed for mitochondrion-encoded protein
genes, whose coding regions are often shortened in the
C-terminal portion compared with their more primitive
counterparts.
Events implicated in changes in the overall organization
of mitochondrial genomes
Changes in mtDNA architecture have been associated
with autonomously replicating mitochondrial plasmids,
which are common in fungi but which also occur in several
plant and protist taxa. The well-characterized mitochon-
drial plasmids in fungi are primarily monomeric linear
molecules with inverted terminal repeats [44]. Insertion of
such linear plasmids into mtDNA probably triggers
conversion of genome conformation from circular mapping
to monomeric linear, as observed in maize cytoplasmic
male sterility [45]. Stable transitions of mtDNA topology
from circular to linear have occurred within relatively
short phylogenetic distances, as seen in yeasts [10], golden
algae [46] and chytridiomycete fungi [43,47].
Changes in mtDNA that lead to size expansion are
mainly caused by an accumulation of repeat sequences and
AT-rich intergenic spacers, mobile elements and introns.
Even within a genus, the length of non-coding intergenic
regions in mtDNA can vary considerably. For example,
mtDNA size in fission yeast species varies between 17 and
$ 80 kbp [48], with multiple arrays of intergenic repeats
accounting for most of the variation. By contrast, rampant
size-expansion of plant mtDNAs seems to have arisen
principally through acquisition of foreign DNA of chlor-
oplast, nuclear, viral and as-yet-unknown origin [49],
involving active DNA uptake [50] with master circles and
subcircles constantly recombining at a low level to create
mixed populations of subgenomic DNA molecules (‘sub-
limons’ [25]). In turn, size reduction of mtDNAs often
occurs by gene transfer to the nucleus (Figure 7); examples
of this phenomenon in flowering plants are especially
notable in terms of both number and frequency [51– 53].
Other strategies of mtDNA size reduction include intron
loss, and elimination of intergenic spacers, sometimes
to such a degree that coding regions of adjacent genes
overlap (e.g. [7,27]).
Rearrangements through insertions, deletions or
recombination follow special rules owing to the multi-
copy character of mitochondrial genomes. In several
http://tigs.trends.com
713
eukaryotic groups, mitochondrial genomes are distributed
over more than one mitochondrion per cell, and the
mtDNA copy number ranges from around ten to several
thousand per cell (often varying with growth conditions).
The multiplicity of mtDNA molecules within the cell
implies that mitochondrial mutations might accumulate
gradually, without immediate deleterious impact. This
could partly explain the accelerated evolutionary rates of
mtDNA relative to nuclear DNA that are primarily
observed in animals (e.g. vertebrates and insects),
fungi (e.g. yeasts and fission yeasts) and certain protists
(e.g. ciliates and slime moulds).
In human somatic cells, it has been well-documented
that mtDNA accumulates point mutations and large
deletions, predominantly in brain and heart muscle of
older individuals (. 40 years) [54]. This high incidence of
mutations has been attributed to several factors, including
slipped mispairing at direct sequence repeats, topoisome-
rase cleavage of mtDNA, the absence of excision repair,
and incomplete repair of damage caused by free-radicals
produced as a by-product of oxidative phosphorylation
[55]. However, as there are few comparative biochemical
studies in other eukaryotes, the specific mechanisms
underlying the particularly high occurrence of mitochon-
drial mutations in human versus other eukaryotes
remains largely unknown.
Exceptional horizontal transfer of mitochondrial genes
Although the evolutionary transfer of genes from organelle
genomes is well-documented (Figure 7), there are few
reports to-date of the transfer of genetic material into the
mitochondrial genome and, especially, between mitochon-
drial genomes. Only in flowering plants (angiosperms)
does incorporation of nuclear and, particularly, chloroplast
DNA sequences into mtDNA occur frequently; such
transfer is not evident among non-angiosperm land plants
or in green algae [49]. Exchange of mobile elements (group
I and group II introns) between mtDNA and non-mtDNA
sources has been inferred on the basis of sequence
comparisons [56– 58], whereas mtDNA-to-mtDNA trans-
fer is suggested by the presence of highly similar introns at
identical positions in Homologous mitochondrial genes of
distantly related species [59 – 61]. A few cases have been
described that involve transfer of coding regions between
mitochondrial genomes; one of these examples involves
fungi (probably from two different classes) and results in a
hybrid atp6 gene [62].
Recently, widespread horizontal transfer of mitochon-
drial genes (both respiratory chain and ribosomal protein)
has been reported to occur between distantly related
angiosperms, even between dicotyledons and monocotyle-
dons [63]. The genomic consequences of these mtDNA-to-
mtDNA transfers include gene duplication, recapture of
genes previously lost through transfer to the nucleus and,
in one particularly notable case, a chimeric, half-monocot –
half-dicot rps11 gene. It is estimated that these transfer
events occurred between 30 and 140 million years ago in
individual cases. It would seem that transfer of genetic
material into angiosperm mtDNA occurs relatively fre-
quently on an evolutionary timescale, regardless of
whether such transfer emanates from the chloroplast
714 Review TRENDS in Genetics Vol.19 No.12 December 2003

(a) Gene export and loss

α-Proteobacterial
endosymbiont Mitochondrion

Primitive Modern
eukaryote eukaryote

(b) Primitive nucleus Nucleus

Acquisition of
symbiont genes and foreign genes;
gene substitutions
TRENDS in Genetics

Figure 7. Fate of proto-mitochondrial genes. Early gene migration has taken two main routes. (a) The a-proteobacterial symbiont underwent massive gene loss during tran-
sition to an organelle. Among the genes essential for mitochondrial functions, the majority was transferred to the nucleus, with a small portion remaining in the evolving
mitochondrial DNA (mtDNA). (b) The hypothetical primitive nucleus of the host [either an amitochondriate eukaryote (see text) [82] or an Archaea-related organism [83])
acquired several hundred symbiont genes, some of which were substituted for nuclear genes. In some instances, nucleus-acquired DNA from other sources replaced a
mitochondrial function initially encoded by the a-proteobacterial symbiont. For example, mtDNA in jakobid flagellates contains genes specifying the components of a typi-
cal eubacteria-like mitochondrial RNA polymerase, whereas in all other eukaryotes, this enzyme has a T3-phage-like structure and is encoded by the nuclear genome [84].
Genes are colour-coded as follows: red, a-proteobacterial genes; blue, nuclear genes of the original host; yellow, a-proteobacterial genes that replaced host genes; green,
foreign genes acquired from sources other than the genome of the mitochondrial ancestor.

genome of the same species or from the mtDNA of a
different plant species. Successful functional transfer of
genes into angiosperm mtDNA might be facilitated by the
expansive, largely non-coding nature of this genome [64],
whose expression seems little constrained by gene location
and overall organization.
Are evolutionary trends similar in mitochondrial and
nuclear genomes?
With the recent availability of nuclear genome data from
several eukaryotes, we are now in a position to compare
evolutionary trends for the mitochondrial and nuclear
genomes of a given organism. One readily measurable
parameter is the proportion of non-coding sequence in the
two genomes. For example, human mtDNA is tightly
packed with intron-less genes [65], whereas human
nuclear genes contain a large number of introns separated
by extensive intergenic regions [66,67]. By contrast, genes
in yeast mtDNA are intron-rich and separated by long non-
coding stretches [68], whereas yeast nuclear DNA har-
bours closely spaced and mostly intron-less genes [69].
Finally, in the pufferfish Fugu, gene density is high in both
genomes [70,71]. Comparison of the relative gene numbers
in the two genomes shows similarly unrelated variations,
implying that the evolutionary trends shaping coding
capacity and density in the nuclear versus mitochondrial
genome proceed independently from one another in a given
organism.
Another issue that can now be investigated is whether
the propensity to insert foreign DNA is the same for the
mitochondrial genome and nuclear genome of a given
organism. In flowering plants, acquisition of foreign DNA
is rampant both in the nucleus and in mitochondria (but
not in chloroplasts), whereas in yeast, DNA escapes from
mitochondria to the nucleus (but not in the opposite
http://tigs.trends.com
direction) at a notably high frequency [72]. Moreover, in
primates, nuclear genomes contain substantial numbers of
mtDNA insertions [73], but the mitochondrial genome in
these cases is free of foreign DNA. These contrasting
examples illustrate the concept that evolution of the
nuclear and organellar genomes of a given eukaryote can
proceed in very different directions.
Does a parasitic lifestyle accelerate mtDNA evolution?
It is tempting to speculate that mtDNA might be under
more relaxed selection in parasitic lineages, leading to the
emergence of exotic mitochondrial genome features, as
described earlier. Indeed, mtDNA with the smallest
number of genes is found in Plasmodium, and the most
extreme case of mRNA editing occurs in trypanosomes,
both of which are obligate parasites. Somewhat surpris-
ingly, this correlation does not withstand careful scrutiny.
For example, Prototheca wickerhamii, a non-photosyn-
thetic chlorophycean alga and (mild) human pathogen, has
one of the most primitive mitochondrial genomes
within this group of green algae [38,74,75]. By
contrast, several free-living, photosynthetic relatives
have highly derived mtDNAs: Scenedesmus obliquus
has an unusual translation code and substantially
fewer mtDNA-encoded genes than P. wickerhamii
mtDNA [38]. In addition, Pedinomonas minor [21]
and Chlamydomonas reinhardtii (and its close rela-
tives) have mtDNAs that are typical cases of reduced
mitochondrial genomes [76]. Other examples of non-
pathogens with highly derived mtDNAs are Euglena
gracilis [77], baker’s and fission yeast (and its close
relatives), animals in general, and Amoebidium para-
siticum [15], a saprotrophic ectosymbiont that is
frequently found on crustaceans and insects that
inhabit fresh water ponds.
 Review TRENDS in Genetics Vol.19 No.12 December 2003
Why do we see the highly derived mtDNAs in some
species but not in others? One possible explanation is that
derived mitochondrial genomes are a consequence of
accelerated growth rates (i.e. they tend to occur in
‘weeds’, that is, organisms with short generation times).
Some such species (e.g. yeasts) might be highly specialized
to thrive in specific environments, whereas other organ-
isms of this type (such as chlamydomonad algae) occur
more widely, dominating over other species through more
efficient propagation.
Mitochondrial DNA variability: implications for gene
expression
In our quest to understand the role of genome organization
in gene expression, a comparative mitochondrial genomics
approach is only the first step. Complete sequencing of
mtDNA enables the identification of genes and permits one
to make inferences about unusual modes of gene
expression (e.g. split and divergently transcribed genes,
or fused ORFs). However, only systematic and compre-
hensive analysis of the transcriptome (the portion of the
genome that is transcribed) will reveal: (i) whether all
identified genes are active, (ii) whether co- or post-
transcriptional processes, such as RNA editing, alter
encoded genetic information on the way to its expression,
and (iii) whether RNA species (perhaps having regulatory
functions) are synthesized in addition to the usual
informational and structural RNAs. Identification of a
gene, even one having all the hallmarks of a functional
unit, is no guarantee that it is active, as exemplified by the
silent cox2 gene in soybean mtDNA [78]. Even where
transcription of a protein-coding gene is shown, definitive
evidence of function ultimately requires identification of
the corresponding protein. This criterion has been
satisfied in very few cases, although direct analysis of
the mitochondrial proteome is rapidly gaining favour.
Concluding remarks
Comparative mitochondrial genomics, especially in pro-
tists, has revealed an often perplexing diversity in overall
organization and mode of expression of mtDNA. The
comparative approach has also amply demonstrated that
this extreme structural divergence co-exists with the
conserved functional role of the mitochondrial genome.
In this respect, mtDNA is an evolutionary playground par
excellence. No correlation is evident in the mode and tempo
of evolutionary change in the mitochondrial and nuclear
genomes across the broad spectrum of eukaryotic lineages.
Acknowledgements
This project was supported by grants from the Canadian Institutes for
Health Research (MSP14226, MOP42475 and MOP4124). Salary and
interaction support from the Canadian Institute for Advanced Research
(G.B., M.W.G. and B.F.L.) and the Canada Research Chairs Programme
(M.W.G. and B.F.L.) is gratefully acknowledged.
References
1 Saraste, M. (1999) Oxidative phosphorylation at the fin de siecle.
Science 283, 1488 – 1493
2 Herrmann, J.M. (2003) Converting bacteria to organelles: evolution of
mitochondrial protein sorting. Trends Microbiol. 11, 74– 79
3 Embley, T.M. et al. (2003) Mitochondria and hydrogenosomes are two
http://tigs.trends.com
715
forms of the same fundamental organelle. Philos. Trans. R. Soc. Lond.
B Biol. Sci. 358, 191 – 201
4 Roger, A.J. and Silberman, J.D. (2002) Cell evolution: mitochondria in
hiding. Nature 418, 827– 829
5 Gray, M.W. et al. (1999) Mitochondrial evolution. Science 283,
1476– 1481
6 Lang, B.F. et al. (1999) Mitochondrial genome evolution and the origin
of eukaryotes. Annu. Rev. Genet. 33, 351 – 397
7 Boore, J.L. (1999) Animal mitochondrial genomes. Nucleic Acids Res.
27, 1767 – 1780
8 Bendich, A.J. (1993) Reaching for the ring: the study of mitochondrial
genome structure. Curr. Genet. 24, 279 – 290
9 Bendich, A.J. (1996) Structural analysis of mitochondrial DNA
molecules from fungi and plants using moving pictures and pulsed-
field gel electrophoresis. J. Mol. Biol. 255, 564 – 588
10 Nosek, J. and Tomaska, L. Mitochondrial genome diversity: evolution
of the molecular architecture and replication strategy. Curr. Genet.
(in press)
11 Burger, G. and Lang, B.F. (2003) Parallels in genome evolution in
mitochondria and bacterial symbionts. IUBMB Life 55, 205 – 212
12 Armstrong, M.R. et al. (2000) A multipartite mitochondrial genome in
the potato cyst nematode Globodera pallida. Genetics 154, 181 – 192
13 Watanabe, K.I. et al. (1999) Mitochondrial genes are found on
minicircle DNA molecules in the mesozoan animal Dicyema. J. Mol.
Biol. 286, 645 – 650
14 Lang, B.F. et al. (2002) The closest unicellular relatives of animals.
Curr. Biol. 12, 1773– 1778
15 Burger, G. et al. (2003) Unique mitochondrial genome architecture in
unicellular relatives of animals. Proc. Natl. Acad. Sci. U. S. A. 100,
892– 897
16 Morris, J.C. et al. (2001) Replication of kinetoplast DNA: an update for
the new millennium. Int. J. Parasitol. 31, 453 – 458
17 Feagin, J.E. (2000) Mitochondrial genome diversity in parasites. Int.
J. Parasitol. 30, 371 – 390
18 Notsu, Y. et al. (2002) The complete sequence of the rice (Oryza sativa
L.) mitochondrial genome: frequent DNA sequence acquisition and
loss during the evolution of flowering plants. Mol. Genet. Genomics
268, 434 – 445
19 Ward, B.L. et al. (1981) The mitochondrial genome is large and
variable in a family of plants (cucurbitaceae). Cell 25, 793 – 803
20 Lang, B.F. et al. (1997) An ancestral mitochondrial DNA resembling a
eubacterial genome in miniature. Nature 387, 493 – 497
21 Turmel, M. et al. (1999) The complete mitochondrial DNA sequences of
Nephroselmis olivacea and Pedinomonas minor. Two radically
different evolutionary patterns within green algae. Plant Cell 11,
1717– 1730
22 Lunt, D.H. et al. (1998) Mitochondrial DNA variable number tandem
repeats (VNTRs): utility and problems in molecular ecology. Molec.
Ecol. 7, 1441– 1455
23 Paquin, B. and Lang, B.F. (1996) The mitochondrial DNA of Allomyces
macrogynus: the complete genomic sequence from an ancestral fungus.
J. Mol. Biol. 255, 688– 701
24 Paquin, B. et al. (2000) Double-hairpin elements in the mitochondrial
DNA of Allomyces: evidence for mobility. Mol. Biol. Evol. 17,
1760– 1768
25 Andre, C. et al. (1992) Small repeated sequences and the structure of
plant mitochondrial genomes. Trends Genet. 8, 128– 132
26 Cummings, D.J. et al. (1990) The complete DNA sequence of the
mitochondrial genome of Podospora anserina. Curr. Genet. 17,
375– 402
27 Burger, G. et al. (1995) The mitochondrial DNA of the amoeboid
protozoon, Acanthamoeba castellanii: complete sequence, gene content
and genome organization. J. Mol. Biol. 245, 522 – 537
28 Ogawa, S. et al. (2000) The mitochondrial DNA of Dictyostelium
discoideum: complete sequence, gene content and genome organiz-
ation. Mol. Gen. Genet. 263, 514– 519
29 Lonergan, K.M. and Gray, M.W. (1996) Expression of a continuous
open reading frame encoding subunits 1 and 2 of cytochrome c oxidase
in the mitochondrial DNA of Acanthamoeba castellanii. J. Mol. Biol.
257, 1019– 1030
30 Hanson, M.R. (1991) Plant mitochondrial mutations and male sterility.
Annu. Rev. Genet. 25, 461 – 486
31 Oudot-Le Secq, M.P. et al. (2001) The complete sequence of a brown
716 Review TRENDS in Genetics Vol.19 No.12 December 2003
algal mitochondrial genome, the ectocarpale Pylaiella littoralis (L.)
Kjellm. J. Mol. Evol. 53, 80 – 88
32 Bullerwell, C.E. et al. (2000) A novel motif for identifying rps3 homologs
in fungal mitochondrial genomes. Trends Biochem. Sci. 25, 363–365
33 Heinonen, T.Y. et al. (1987) Rearranged coding segments, separated by
a transfer RNA gene, specify the two parts of a discontinuous large
subunit ribosomal RNA in Tetrahymena pyriformis mitochondria.
J. Biol. Chem. 262, 2879– 2887
34 Schnare, M. et al. (1986) A discontinuous subunit ribosomal RNA in
Tetrahymena pyriformis mitochondria. J. Biol. Chem. 261, 5187– 5193
35 Boer, P.H. and Gray, M.W. (1988) Scrambled ribosomal RNA gene
pieces in Chlamydomonas reinhardtii mitochondrial DNA. Cell 55,
399 – 411
36 Gillespie, D.E. et al. (1999) The fragmented mitochondrial ribosomal
RNAs of Plasmodium falciparum have short A tails. Nucleic Acids Res.
27, 2416 – 2422
37 Edqvist, J. et al. (2000) Expression of mitochondrial protein-coding
genes in Tetrahymena pyriformis. J. Mol. Biol. 297, 381– 393
38 Nedelcu, A.M. et al. (2000) The complete mitochondrial DNA sequence of
Scenedesmus obliquus reflects an intermediate stage in the evolution of
the green algal mitochondrial genome. Genome Res. 10, 819–831
39 Perez-Martinez, X. et al. (2001) Subunit II of cytochrome c oxidase in
chlamydomonad algae is a heterodimer encoded by two independent
nuclear genes. J. Biol. Chem. 276, 11302 – 11309
40 Adams, K.L. et al. (2001) Mitochondrial gene transfer in pieces: fission
of the ribosomal protein gene rpl2 and partial or complete gene
transfer to the nucleus. Mol. Biol. Evol. 18, 2289– 2297
41 Gray, M.W. et al. (1998) Genome structure and gene content in protist
mitochondrial DNAs. Nucleic Acids Res. 26, 865– 878
42 Seif, E.R. et al. (2003) Mitochondrial RNase P RNAs in ascomycete
fungi: lineage-specific variations in RNA secondary structure. RNA 9,
1073 – 1083
43 Bullerwell, C.E. et al. (2003) Evolution of monoblepharidalean fungi
based on complete mitochondrial genome sequences. Nucleic Acids
Res. 31, 1614– 1623
44 Kennell, J.C. and Cohen, S.M. In Handbook of Fungal Biotechnology
(Arora, D., ed.), Marcel Dekker (in press)
45 Schardl, C.L. et al. (1985) Mitochondrial DNA rearrangements associ-
ated with fertile revertants of S-type male-sterile maize. Cell 43,
361–368
46 Chesnick, J.M. et al. (2000) The mitochondrial genome of the
stramenopile alga Chrysodidymus synuroideus. Complete sequence,
gene content and genome organization. Nucleic Acids Res. 28,
2512–2518
47 Forget, L. et al. (2002) Hyaloraphidium curvatum: a linear mitochon-
drial genome, tRNA editing, and an evolutionary link to lower fungi.
Mol. Biol. Evol. 19, 310 – 319
48 Bullerwell, C.E. et al. (2003) A comparison of three fission yeast
mitochondrial genomes. Nucleic Acids Res. 31, 759 – 768
49 Marienfeld, J. et al. (1999) The mitochondrial genome of Arabidopsis is
composed of both native and immigrant information. Trends Plant Sci.
4, 495 – 502
50 Koulintchenko, M. et al. (2003) Plant mitochondria actively import DNA
via the permeability transition pore complex. EMBO J. 22, 1245–1254
51 Adams, K.L. et al. (2000) Repeated, recent and diverse transfers of a
mitochondrial gene to the nucleus in flowering plants. Nature 408,
354 – 357
52 Adams, K.L. et al. (2001) Multiple losses and transfers to the nucleus of
two mitochondrial succinate dehydrogenase genes during angiosperm
evolution. Genetics 158, 1289 – 1300
53 Adams, K.L. et al. (2002) Punctuated evolution of mitochondrial gene
content: high and variable rates of mitochondrial gene loss and
transfer to the nucleus during angiosperm evolution. Proc. Natl. Acad.
Sci. U. S. A. 99, 9905 – 9912
54 Chomyn, A. and Attardi, G. (2003) MtDNA mutations in aging and
apoptosis. Biochem. Biophys. Res. Commun. 304, 519 – 529
55 Wei, Y.H. and Lee, H.C. (2002) Oxidative stress, mitochondrial DNA
mutation, and impairment of antioxidant enzymes in aging. Exp. Biol.
Med. (Maywood) 227, 671 – 682
56 Lonergan, K.M. and Gray, M.W. (1994) The ribosomal RNA gene region
in Acanthamoeba castellanii mitochondrial DNA. A case of evolution-
ary transfer of introns between mitochondria and plastids? J. Mol.
Biol. 239, 476– 499
http://tigs.trends.com
View publication stats
57 Turmel, M. et al. (1995) Evolutionary transfer of ORF-containing
group I introns between different subcellular compartments (chlor-
oplast and mitochondrion). Mol. Biol. Evol. 12, 533 – 545
58 Burger, G. et al. (1999) Complete sequence of the mitochondrial DNA of
the red alga Porphyra purpurea. Cyanobacterial introns and shared
ancestry of red and green algae. Plant Cell 11, 1675– 1694
59 Lang, B.F. (1984) The mitochondrial genome of the fission yeast
Schizosaccharomyces pombe: highly homologous introns are inserted
at the same position of the otherwise less conserved cox1 genes in
Schizosaccharomyces pombe and Aspergillus nidulans. EMBO J. 3,
2129–2136
60 Cho, Y. et al. (1998) Explosive invasion of plant mitochondria by a
group I intron. Proc. Natl. Acad. Sci. U. S. A. 95, 14244 – 14249
61 Cho, Y. and Palmer, J.D. (1999) Multiple acquisitions via horizontal
transfer of a group I intron in the mitochondrial cox1 gene during
evolution of the Araceae family. Mol. Biol. Evol. 16, 1155 – 1165
62 Paquin, B. et al. (1994) Interspecific transfer of mitochondrial genes in
fungi and creation of a homologous hybrid gene. Proc. Natl. Acad. Sci.
U. S. A. 91, 11807 – 11810
63 Bergthorsson, U. et al. (2003) Widespread horizontal transfer of
mitochondrial genes in flowering plants. Nature 424, 197 – 201
64 Hanson, M.R. and Folkerts, O. (1992) Structure and function of the
higher plant mitochondrial genome. Int. J. Cytol. 141, 129– 172
65 Anderson, S. et al. (1981) Sequence and organization of the human
mitochondrial genome. Nature 290, 457– 465
66 Lander, E.S. et al. (2001) Initial sequencing and analysis of the human
genome. Nature 409, 860 – 921
67 Venter, J.C. et al. (2001) The sequence of the human genome. Science
291, 1304– 1351
68 Foury, F. et al. (1998) The complete sequence of the mitochondrial
genome of Saccharomyces cerevisiae. FEBS Lett. 440, 325 – 331
69 Giaever, G. et al. (2002) Functional profiling of the Saccharomyces
cerevisiae genome. Nature 418, 387 – 391
70 Elmerot, C. et al. (2002) The mitochondrial genome of the pufferfish,
Fugu rubripes, and ordinal teleostean relationships. Gene 295,
163– 172
71 Aparicio, S. et al. (2002) Whole-genome shotgun assembly and analysis
of the genome of Fugu rubripes. Science 297, 1301– 1310
72 Thorsness, P.E. and Fox, T.D. (1990) Escape of DNA from mitochondria
to the nucleus in Saccharomyces cerevisiae. Nature 346, 376 – 379
73 Woischnik, M. and Moraes, C.T. (2002) Pattern of organization of
human mitochondrial pseudogenes in the nuclear genome. Genome
Res. 12, 885 – 893
74 Wolff, G. et al. (1994) Complete sequence of the mitochondrial DNA of
the chlorophyte alga Prototheca wickerhamii. Gene content and
genome organization. J. Mol. Biol. 237, 75 – 86
75 Kuck, U. et al. (2000) DNA sequence analysis of the complete
mitochondrial genome of the green alga Scenedesmu obliquus:
evidence for UAG being a leucine and UCA being a non-sense codon.
Gene 253, 13 – 18
76 Gray, M.W. and Boer, P.H. (1988) Organization and expression of algal
(Chlamydomonas reinhardtii) mitochondrial DNA. Philos. Trans.
R. Soc. Lond. B Biol. Sci. 319, 135 – 147
77 Tessier, L.H. et al. (1997) The cox1 gene from Euglena gracilis: a protist
mitochondrial gene without introns and genetic code modifications.
Curr. Genet. 31, 208 – 213
78 Covello, P.S. and Gray, M.W. (1992) Silent mitochondrial and active
nuclear genes for subunit 2 of cytochrome c oxidase (cox2) in soybean:
evidence for RNA-mediated gene transfer. EMBO J. 11, 3815 – 3820
79 Gabaldon, T. and Huynen, M.A. (2003) Reconstruction of the proto-
mitochondrial metabolism. Science 301, 609
80 Rasmusson, A.G. et al. (1999) Homologues of yeast and bacterial
rotenone-insensitive NADH dehydrogenases in higher eukaryotes:
two enzymes are present in potato mitochondria. Plant J. 20, 79 – 87
81 Baldauf, S.L. (2003) The deep roots of eukaryotes. Science 300,
1703– 1706
82 Cavalier-Smith, T. (1983) A 6-kingdom classification and a unified
phylogeny. In Endocytobiology II (Schwemmler, W. and Schenk,
H.E.A., eds), pp. 1027– 1034, De Gruyter
83 Martin, W. and Müller, M. (1998) The hydrogen hypothesis for the first
eukaryote. Nature 392, 37 – 41
84 Cermakian, N. et al. (1997) On the evolution of the single-subunit RNA
polymerases. J. Mol. Evol. 45, 671 – 681