A B S T RAe T
Gene or genome duplication events increase the amount of genetic material available to increase the genomic, and thereby phenotypic, complexity of organisms during evolution. Gene duplication and transfer events have been important to molecular evolution in all three domains of life, and may be the first step in the emergence of new gene functions. Gene transfer events have been proposed as another accelerator of evolution. The duplicated gene or genome, mainly nuclear, has been the subject of several recent reviews. In addition to the nuclear genome, organisms have organelle genornes, including mitochondrial genome. In this review, we briefly summarize gene duplication and transfer events in the plant mitochondrial genome.
Mitochondria are membrane-enclosed organelles that occur in most eukaryotic cells. Mitochondria have inner and outer membranes composed of phospholipid bilayers and proteins . Mitochondria, which have been called "cellular power plants", produce most of the cell's supply of adenosine triphosphate (ATP), which is used as a source of energy. In addition to supplying energy to the cell, mitochondria are involved in a range of processes, such as signaling, cellular differentiation, and cell death, as well as control of the cell cycle and cell growth . Mitochondria are not necessarily inherited solely through the maternal line; they can be inherited from both parents .
The non-Mendelian genetics of extracellular genomes were first reported a century ago. Mitochondria are believed to be the products of endosymbiotic events. Most of the DNA present in eukaryotic organisms is in the cell nucleus, but they also have independent mitochondrial genomes [4-5]. Mitochondrial DNA is maternally inherited in most multicellular organisms. Coding regions in the mitochondrial genome accumulate sequence changes very slowly, but the linear arrangement of genes changes quite quickly . Mitochondrial DNA has direct repeats spread throughout the genome. More than 1000 complete mitochondrial DNA sequences have been
published for organisms including mammals, protists, ascomycete fungi and plants [7-10].
Gene duplication events as the primary driving force for evolution
Gene duplication and subsequent divergence are important in the evolution of genes by providing genetic material from which novel functions can arise [11-13]. The function of genes after duplication can be categorized as neofunctionalization, subfunctionalization. or nonfunctionalization . Careful analysis of duplicated regions shows that the majority of duplicated genes disappear during evolution . Some duplicated genes may be lost by accumulation of deleterious mutations (nonfunctionalization). Paralogous genes (newly duplicated genes) that are not silenced may be maintained by subfunctionalization (partitioning ancestral functions, with the duplicated genes performing different aspects of the original gene's function) and/or neofunctionalization (one of the genes acquires a novel function) [14.16-18].
There are several pathways by which genes can duplicate [19-22]. Gene or genome duplication events can involve a single gene, a segment of the genome, a single chromosome or even the whole genome. Genome duplications differ from single gene duplications in that whole chromosomes are simultaneously doubled, and the overall number of genes is thereby increased [19,21.22,24]. It has long been known that new genes frequently emerge through gene and genome duplication. Spontaneous duplication of large chromosomal segments has been demonstrated experimentally in yeast [25-26]. Polyploidy, which results in duplication of the whole gene complement of an organism, is
widespread in eukaryotes, and is probably one of the main mechanisms underlying evolutionary divergence [27-28].
The mitochondrion is a nearly autonomous organelle that contains the biochemical machinery necessary to replicate and transcribe its own genome and to synthesize protein. In addition to the nuclear genome, organisms have mitochondrial genomes, most of which undergo intra- and intergenomic recombination and rearrangement . Mitochondrial genomes are simplified relics of the much larger cellular genomes of their cyanobacterial and proteobacterial ancestors . Gene duplication is an important process in mitochondrial evolution, but duplication of large fragments of mitochondrial genomes is infrequent. Knowledge of the sequences of the mitochondrial genome of a number of organisms has made it possible to conduct comprehensive searches for duplicated genes, enabling informative study of their evolution . The availability of complete mitochondrial and nuclear genome sequences has made it possible to study the extent of gene duplication events and gene transfer events.
Gene duplication events in plant mitochondria
The plant mitochondrial genome is a circular double-stranded DNA molecule that encodes tRNAs, rRNAs, ribosomal proteins, and a portion of the enzymes used in respiration [32-33]. Plant mitochondrial genomes range in size from 200 kb to 2400 kb and are at least 10-100 times larger than animal mitochondrial genomes [34-35]. At 208 kb, Brassica hirta is one of the smallest and Cucumis rneio at 2300 kb is the largest known mitochondrial genome in higher plants [6,34,36]. The plant mitochondrial genome has an unusually dynamic structure due to recombination between repeated sequences, which generates a population of molecules of different sizes and molecular configurations. The plant mitochondrial genome has a very low substitution rate, and its evolution is characterized by frequent structural rearrangements [36-37]. Several descriptive models have been proposed to explain the occurrence of deletion-duplication events in the plant mitochondrial genome [37-39].
The entire mitochondrial genome of rapeseed (Brassica napus L.), which contains a 2427 bp sequence as a direct repeat, was sequenced by Handa . This sequence includes the first exon, the intron, and part of the second exon of the cox2 gene. Due to duplication, two copies of cox2 genes exist in rapeseed, although these copies diverge from each other 55 bp upstream of the stop codon. One copy (cox2-1) is homologous to other plant mitochondrial cox2 genes, but the other copy (cox2-2) has an extension that shows no homology to any other sequence examined to date .
The cucumber (Cucumis sativus) has some unique attributes that make it a potential model system for mitochondrial transformation of higher plants. Microspores have relatively few, huge mitochondria, which have paternal transmission. The cucumber has unique mitochondrial mutations that result in strongly mosaic phenotypes . Lilly and Havey investigated mitochondrial genome expansion within the cucurbits using hybridization to select mitochondrial sequences present in high copy numbers in cucumber and at low levels in watermelon (Citrullus lanatus). Lilly and Havey sequenced 15 clones to identify sequences repeated throughout the cucumber mitochondrial genome. On the basis of dot-blot hybridizations, seven repetitive DNA motifs account for over 13% of the cucumber mitochondrial genome, equaling over 50% of the size of the Arabidopsis mitochondrial genome. Sequence analysis of136 kb of cucumber mitochondrial DNA revealed only 11.2% with significant homology to previously characterized mitochondrial sequences, 2.4% to chloroplast DNA, and 15% to the seven repetitive DNA motifs. The remaining 71.4% of the sequence was unique to the cucumber
mitochondrial genome. These results demonstrate that the expanded cucumber mitochondrial genome is due, in part, to extensive duplication of short repetitive sequences, possibly by recombination and/or replication slippage .
There are two divergent copies of the chloroplast origin rps13 gene, which encodes ribosomal protein S13, are found in the nucleus of the rosids Arabidopsis, Gossypium, and Glycine. One is nucp-rpsI3, which encodes chloroplast-imported RPS13; the other is numit-rpsI3, which encodes mitochondria-imported RPS13 . The function of numit-rpsl3 has been modified after gene duplication, and one could argue that numit-rpsl3 has gained a new function. Subsequently mt-rpsl3 was lost from mitochondrial DNA many times during the evolutionary history ofrosids. Those organellar rps13 genes in rosids provide a distinctive case of gene duplication involving the co-evolution of the nuclear and cytoplasmic genomes [41-42].
Gene transfer events between mitochondrial and nuclear genomes
In addition to the nuclear genome, plants have chloroplast and mitochondrial genomes, all of which undergo intra- and intergenomic recombination and rearrangement. The nuclear, chloroplast and mitochondrial genomes have been sequenced in some plants, such as Arabidopsis [32,43,44] and rice [9,45,46]. With the completion of those plant genome sequencing projects, it was possible to identify transfer events from the organelles to the nucleus on a whole-genome scale.
It is well known that mitochondria donated many genes to nuclear chromosomes during evolution. Rujan and Martin compared 3961 Arabidopsis nuclear protein-coding genes with the complete set of proteins from yeast and 17 reference prokaryotic genomes . The degree of conservation in protein sequences in addition to lateral gene transfer between free-living prokaryotes pose substantial challenges to genome phylogenetics.
To date, several genes transferred from the mitochondrial genome to the nuclear genome have been identified in flowering plants. However, the mechanism of gene transfer events is only poorly understood . Gene transfer events between the mitochondrial genome and the nuclear genome in plants are believed to often occur as single gene transfers through an RNA intermediate. The majority of mitochondria gene transfer events have been reported to range from hundreds to thousands of base pairs . The sequence analysis of Arabidopsis thaliana chromosome 2 revealed a mitochondrial-to-nuclear DNA transfer of nearly the entire mitochondrial genome into the pericentric region on the short arm. The size of the mitochondrial DNA insert was about 270 kb . However, DNA fiber-based fluorescence in situ hybridization analysis revealed that the mitochondrial DNA insert is about 620 kb, which is near the centromere on chromosome 2 .
A promiscuous nuclear sequence containing a mitochondrial DNA fragment was isolated from rice by Kubo and his colleagues , who found that the integration ofthe mitochondrial sequence into the nuclear genome was mediated by a DNA fragment, and that the nuclear sequence was transcribed and spliced, but it appeared to be a pseudogene. The mitochondrial sequence was integrated in an antisense orientation into the pre-existing V-ATPase B pseudogene, which can be transcribed and spliced. They suggested that the DNA transfer event may be a case of unsuccessful gene transfer from mitochondrion to nucleus (Fig. 1).
The rps13 gene, encoding mitochondrial ribosomal protein S13, is normally present in the mitochondrial genome of higher plants, but is lacking from the A. thaliana mitochondrial genome [54-55]. Mollier et al. demonstrated that the nuclear gene encoding the plastid S13 has been partially duplicated in A. thaliana, such that
the copy has lost the exon encoding the plastid transit peptide and has acquired a sequence capable of encoding a mitochondrial targeting sequence. The mitochondrial S13 ribosomal protein has probably been replaced by its homologue from plastids in A. thaliana. The differences between the S13 gene sequence of the mitochondrion and the chloroplast suggest that the gene duplication occurred after the Brassicaceae arose but before the divergence of Arabidopsis and Brassica .
The complete mitochondrial genome of rice has been sequenced. Notsu et al. found that 6.3% and 13.4% of the mitochondrial genome sequence is derived from the plastid and the nuclear genome, respectively . They demonstrated frequent and independent DNA sequence flow among the mitochondrial, plastid and nuclear genomes during the evolution of flowering plants, and this may account for the range of genetic variation observed between the mitochondrial genomes of higher plants .
Conclusion and perspective
The complete genome sequences, including the mitochondrial and nuclear genomes, of more and more organisms are becoming available, and this can be considered a major step forward toward exploiting the usefulness of mitochondrial genetic engineering technology. Earlier work showed that many gene transfers to the nuclear occurred during mitochondrial evolution, but there is no reliable estimate of the total number of genes that have been transferred.
Gene duplication is a fundamental process in the evolution of eukaryotic genomes. After duplication, one copy of a gene may undergo divergence in sequence, expression pattern, and function, and the rate of gene duplication is an important parameter in the study of evolution. During the past decade, the amount of sequence data (primarily DNA sequence data) has increased nearly lOO-fold, and will continue to increase rapidly as the result of immense technical progress in DNA sequencing. Completely sequenced mitochondrial genomes are a valuable source of data for determining the evolutionary history of the organelle. Many mitochondrial enzymatic subunits are nuclear-encoded, cytoplasmically translated, and imported into the mitochondria. Gene duplication events in the mitochondrial genome and gene transfer events between the mitochondrial genome and the nuclear genome are important processes in the evolution ofthe eukaryotic cell.
Authors acknowledge the funding from Project for International Scientific and Technological Cooperation (Shanghai China-Alberta Canada); Shanghai Rising-Star Program (08QH14021); Hi-tech research and development program of China (2006AA10Zl17; 2008AA10Z401). We are grateful to Prof. Max Cheng (Department of Plant Sciences, University of Tennessee, USA) for his useful comments on the manuscript
Gene duplication Evolution