W. Martin, "Molecular evolution: Lateral gene transfer and other possibilities" (2005)

"Heredity" June 2005, Volume 94, Number 6, Pages 565-566; http://www.nature.com/cgi-taf/DynaPage.taf?file=/hdy/journal/v94/n6/full/6800659a.html

Molecular evolution: Lateral gene transfer and other
possibilities

W Martin1

1The Institute of Botany, University of Düsseldorf, Germany
Correspondence to: W Martin, e-mail: [email protected]

Heredity (2005) 94, 565-566. doi:10.1038/sj.hdy.6800659
Published online 30 March 2005

                      Two recent reports suggesting that extensive lateral gene transfer occurs
                      among higher plants clash with our view of evolution as Darwin understood
                      it.
                      The concept of descent with modification has proven exquisitely robust, with
                      only two genuine mechanistic additions to Darwin's principles of natural
                      selection operating on variation among progeny, having emerged over the
                      last 150 years. One is endosymbiosis, where highly divergent lineages merge
                      outright, such as the origin of chloroplasts from cyanobacteria or the origin
                      of mitochondria from proteobacteria. The other is lateral, or horizontal, gene
                      transfer (LGT), where disparate lineages occasionally exchange parts of
                      their genetic fabric. Genome sequences have provided sound evidence that
                      both endosymbiosis among eukaryotes, and LGT - among prokaryotes -
                      are indeed real, although there is still much debate as to just how frequently
                      either has occurred during evolution. That debate now continues.
                      In 2003, Bergthorsson's team reported sequences homologous to plant
                      mitochondrial DNA (mtDNA) from a variety of species, obtained using
                      polymerase chain reaction (PCR) with conserved primers against
                      protein-coding regions of plant mtDNA. As some sequences in the
                      phylogenies branched in very unusual positions, the authors concluded that
                      frequent lateral transfer of mitochondrial DNA between distantly related
                      plants had caused this pattern. Viruses, bacteria, fungi, insects, pollen, even
                      meteorites and grafting were suggested as vectors for this exchange
                      (Bergthorsson et al, 2003). The authors provided a figure showing genes
                      being laterally transferred not only between species but also from ancient
                      lineages in the past to more recent lineages, interpretations that 'imply the
                      existence of the transferred gene in an intermediate, unidentified
                      vectoring agent or host plant for millions of years' (Bergthorsson et al,
                      2003) - yet without mutation, one should add. Several of the unsually
                      branching sequences involved the shrubby flowering plant Amborella
                      trichopoda, prompting a more extensive search among DNA samples from
                      this species.
                      That follow-up study on Amborella has now appeared (Bergthorsson et al,
                      2004) and is no less eyebrow-raising. Conserved primers were designed for
                      the 31 protein-coding genes typical of higher plant mtDNA. In total, 20 of
                      the 31 primer pairs generated two or more different PCR amplification
                      products with Amborella DNA as the substrate and these mutiple products
                      branched in disparate parts of the trees. Bergthorsson et al (2004)
                      interpreted this as evidence for 'massive' LGT from a myriad of higher plant
                      donors, and concluded that Amborella mtDNA 'has sustained
                      proportionately more HGT than any other eukaryotic, or perhaps even
                      prokaryotic, genome yet examined'. If true, this would be an
                      unprecedented situation for three reasons. First, plant mitochondrial genome
                      sequences have not yet provided evidence for the acquisition of genes from
                      other species (Bergthorsson et al, 2004). Second, the Amborella
                      chloroplast genome sequence reveals no acquisitions from other species
                      (Goremykin et al, 2003). Third, no reports of widespread lateral acquisition
                      from various higher plant donors have emerged from any plant nuclear
                      genome sequenced so far. Is there really something special about
                      Amborella that makes it an LGT-haven? Is higher plant mtDNA hopping
                      among species faster than we can sequence it? Or are there possible
                      alternative interpretations of the observations other than LGT?
                      If we exclude DNA contamination from other sources as a possible factor in
                      their results, as Bergthorsson's team (2003, 2004) did, we can still ask what
                      positive evidence there is that the sequences in question are indeed
                      incorporated in the Amborella mtDNA or nuclear genome to substantiate
                      the LGT case. Specific hybridisation and cloning of the noncoding regions
                      flanking the amplified sequences would have offered the opportunity to see
                      how and where the LGT candidate sequences were integrated in
                      unequivocally endogenous Amborella DNA. Flanking sequences as specific
                      probes are needed in the case of plant mtDNA because it evolves at an
                      inexplicably slow rate: across the deepest comparisons of flowering plants,
                      sequences of mtDNA coding regions are typically 95% identical or more at
                      the nucleotide level and hence will crosshybridise. However, these studies
                      only reported the conserved reading frame sequences, without any flanking
                      regions or integration sites, leaving their chromosomal and intracellular
                      location - mitochondrion or nucleus - open, although Bergthorsson and
                      team (2004) favour a mitochondrial localisation. This could have been
                      clarified by a complete mitochondrial genome sequence for Amborella,
                      contiguous linkage between the LGT candidates and bona fide mtDNA
                      isolated from organelles, or in situ hybridisation with specific probes.
                      Indeed, the recently published mitochondrial genome sequence for tobacco
                      (Sugiyama et al, 2005) revealed that several genes previously reported to
                      have been lost from that genome and transferred to the nucleus are in fact
                      present in the mtDNA, underscoring the value of complete genome data in
                      assessing subcellular gene localisation.
                      There is also the possibility that the unusual PCR products from Amborella
                      stem from substrates somewhere in its genome, but that the unusual trees are
                      not due to LGT. Those who study animal mitochondrial DNA have long
                      known that nuclear pseudogenes of mitochondrial DNA often have unusual
                      sequences (Bensasson et al, 2001). Such nuclear pseudogenes of mtDNA
                      are called 'numts' and have been found in large numbers in most sequenced
                      eukaryotic genomes, particularly among plants (Richly and Leister, 2004).
                      Thalmann et al (2004) recently showed that numts in primates are a serious
                      problem because they are readily, sometimes even preferentially, amplified
                      over genuine mtDNA, even though the organelle copy is present in larger
                      template numbers. Numts are extremely difficult to distinguish from bona
                      fide mtDNA (Bensasson et al, 2001; Thalmann et al, 2004, 2005) and can
                      produce very unusual branching patterns in phylogenetic trees causing
                      primate species to apparently intermingle (Thalmann et al, 2004) in a
                      pattern that would suggest rampant LGT, were the identity of the bona fide
                      mtDNA and the numt not known.
                      The levels of sequence differences between higher plant mtDNA from
                      different orders are low, less than that observed between human and chimp
                      mtDNAs. For example, the rps2 sequences representing the deeply
                      diverging dicot orders Laurales and Magnoliales - separated by roughly
                      150 million years (Bergthorsson et al, 2003) - have only two nucleotide
                      differences across 474 sites: G T transversions at positions 89 and 263.
                      The 1.2 kb-long atp1 sequences from the gymnosperm Ginkgo and the
                      angiosperm Illicium, separated by about 300 million years (Kim et al,
                      2004), have only 6% nucleotide differences (Bergthorsson et al, 2004). By
                      comparison, human and chimp sequences for the 1.8-kb long nad5 gene
                      from mtDNA have nearly twice as much (11%) differences. The extremely
                      low rate of substitution in higher plant mtDNA makes it amenable to using
                      conserved primers (Bergthorsson et al, 2003, 2004), but low numbers of
                      substitutions alone does not distinguish whether the PCR products obtained
                      are mtDNA or numts (Bensasson et al, 2001; Thalmann et al, 2004).
                      Adding to these uncertainties is the issue of RNA editing, the C T
                      changes of which affect over 20 codons each in 10 different reading frames
                      of Arabidopsis mtDNA (Geige and Brennicke, 1999). Little is known
                      about the prevalence and among-gene distribution of mitochondrial editing in
                      other higher plant lineages. Other reports for plant-to-plant LGT have also
                      appeared recently (Won and Renner, 2003) but they, too, involved
                      exclusively mtDNA and very small numbers of remarkably distributed
                      sequence differences. There is also the issue of current phylogenetic
                      methods themselves, which are anything but error-free (Holland et al,
                      2004).
                      None of this is to say that the mechanisms and amounts of LGT inferred in
                      the recent findings from plant mtDNA cannot be true. However, the
                      inferences of LGT via meterorites, LGT from the past to the present, and
                      more frequent LGT among shrubs than among prokaryotes are rather
                      surprising. It is prudent, therefore, to consider possible alternative
                      explanations. After all, dinosaur bone DNA once caused quite a stir, but it
                      turned out to be a numt (Zischler et al, 1995). Thus, it will be of interest to
                      see this new LGT evidence corroborated by independent experimental
                      approaches that circumvent PCR and to see its biological significance for the
                      process of heredity among the organisms in question.

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