Journal of Molecular Evolution

, Volume 70, Issue 5, pp 479–491 | Cite as

Testing for Selection on Synonymous Sites in Plant Mitochondrial DNA: The Role of Codon Bias and RNA Editing



Since plant mitochondrial genomes exhibit some of the slowest known synonymous substitution rates, it is generally believed that they experience exceptionally low mutation rates. However, the use of synonymous substitution rates to infer mutation rates depends on the implicit assumption that synonymous sites are evolving neutrally (or nearly so). To assess the validity of this assumption in plant mitochondrial genomes, we examined coding sequence for footprints of selection acting at synonymous sites. We found that synonymous sites exhibit an AT rich and pyrimidine skewed nucleotide composition compared to both non-synonymous sites and non-coding regions. We also found some evidence for selection associated with both biased codon usage and conservation of regulatory sequences involved in mRNA processing, although some of these findings are subject to alternative non-adaptive interpretations. Regardless, the inferred strength of selection appears too weak to account for the variation in substitution rates between the mitochondrial genomes of plants and other multicellular eukaryotes. Therefore, these results are consistent with the interpretation that plant mitochondrial genomes experience a substantially lower mutation rate rather than increased functional constraints acting on synonymous sites. Nevertheless, there are important nucleotide composition patterns (particularly the differences between synonymous sites and non-coding DNA) that remain largely unexplained.


Codon usage bias Mitochondrial genome mtDNA Mutation bias RNA editing Substitution rate Synonymous sites 



We would like to thank Janis Antonovics, Stefan Bekiranov, Lei Li and Martin Wu for helpful discussion of our results. This study was supported by a grant from the NSF (DEB-0808452).

Supplementary material

239_2010_9346_MOESM1_ESM.xls (18 kb)
Supplementary material 1 (XLS 18 kb)
239_2010_9346_MOESM2_ESM.xls (83 kb)
Supplementary material 2 (XLS 83 kb)
239_2010_9346_MOESM3_ESM.xls (74 kb)
Supplementary material 3 (XLS 74 kb)
239_2010_9346_MOESM4_ESM.pdf (104 kb)
Supplementary material 4 (PDF 105 kb)
239_2010_9346_MOESM5_ESM.pdf (167 kb)
Supplementary material 5 (PDF 167 kb)


  1. Adamo A, Pinney JW, Kunova A, Westhead DR, Meyer P (2008) Heat stress enhances the accumulation of polyadenylated mitochondrial transcripts in Arabidopsis thaliana. PLoS One 3:e2889CrossRefPubMedGoogle Scholar
  2. Andolfatto P (2005) Adaptive evolution of non-coding DNA in Drosophila. Nature 437:1149–1152CrossRefPubMedGoogle Scholar
  3. Arabidopsis Genome Initiative (2000) Analysis of the genome sequence of the flowering plant Arabidopsis thaliana. Nature 408:796–815CrossRefGoogle Scholar
  4. Bakker FT, Breman F, Merckx V (2006) DNA sequence evolution in fast evolving mitochondrial DNA nad1 exons in Geraniaceae and Plantaginaceae. Taxon 55:887–896CrossRefGoogle Scholar
  5. Barr CM, Keller SR, Ingvarsson PK, Sloan DB, Taylor DR (2007) Variation in mutation rate and polymorphism among mitochondrial genes in Silene vulgaris. Mol Biol Evol 24:1783–1791CrossRefPubMedGoogle Scholar
  6. Chamary JV, Parmley JL, Hurst LD (2006) Hearing silence: non-neutral evolution at synonymous sites in mammals. Nat Rev Genet 7:98–108CrossRefPubMedGoogle Scholar
  7. Chateigner-Boutin AL, Ramos-Vega M, Guevara-Garcia A, Andres C, de la Luz Gutierrez-Nava M, Cantero A, Delannoy E, Jimenez LF, Lurin C, Small I, Leon P (2008) CLB19, a pentatricopeptide repeat protein required for editing of rpoA and clpP chloroplast transcripts. Plant J 56:590–602CrossRefPubMedGoogle Scholar
  8. Chaw SM, Shih AC, Wang D, Wu YW, Liu SM, Chou TY (2008) The mitochondrial genome of the gymnosperm Cycas taitungensis contains a novel family of short interspersed elements, Bpu sequences, and abundant RNA editing sites. Mol Biol Evol 25:603–615CrossRefPubMedGoogle Scholar
  9. Cho Y, Mower JP, Qiu YL, Palmer JD (2004) Mitochondrial substitution rates are extraordinarily elevated and variable in a genus of flowering plants. Proc Natl Acad Sci USA 101:17741–17746CrossRefPubMedGoogle Scholar
  10. Choury D, Farre JC, Jordana X, Araya A (2004) Different patterns in the recognition of editing sites in plant mitochondria. Nucleic Acids Res 32:6397–6406CrossRefPubMedGoogle Scholar
  11. Crick FH (1966) Codon–anticodon pairing: the wobble hypothesis. J Mol Biol 19:548–555CrossRefPubMedGoogle Scholar
  12. Denver DR, Morris K, Lynch M, Thomas WK (2004) High mutation rate and predominance of insertions in the Caenorhabditis elegans nuclear genome. Nature 430:679–682CrossRefPubMedGoogle Scholar
  13. Drouin G, Daoud H, Xia J (2008) Relative rates of synonymous substitutions in the mitochondrial, chloroplast and nuclear genomes of seed plants. Mol Phylogenet Evol 49:827–831CrossRefPubMedGoogle Scholar
  14. Duret L (2002) Evolution of synonymous codon usage in metazoans. Curr Opin Genet Dev 12:640–649CrossRefPubMedGoogle Scholar
  15. Farre JC, Leon G, Jordana X, Araya A (2001) cis Recognition elements in plant mitochondrion RNA editing. Mol Cell Biol 21:6731–6737CrossRefPubMedGoogle Scholar
  16. Giege P, Brennicke A (1999) RNA editing in Arabidopsis mitochondria effects 441 C to U changes in ORFs. Proc Natl Acad Sci USA 96:15324–15329CrossRefPubMedGoogle Scholar
  17. Glover KE, Spencer DF, Gray MW (2001) Identification and structural characterization of nucleus-encoded transfer RNAs imported into wheat mitochondria. J Biol Chem 276:639–648CrossRefPubMedGoogle Scholar
  18. Grantham R, Gautier C, Gouy M, Jacobzone M, Mercier R (1981) Codon catalog usage is a genome strategy modulated for gene expressivity. Nucleic Acids Res 9:r43–r74CrossRefPubMedGoogle Scholar
  19. Grewe F, Viehoever P, Weisshaar B, Knoop V (2009) A trans-splicing group I intron and tRNA-hyperediting in the mitochondrial genome of the lycophyte Isoetes engelmannii. Nucleic Acids Res 37:5093–5104CrossRefPubMedGoogle Scholar
  20. Hammani K, Okuda K, Tanz SK, Chateigner-Boutin AL, Shikanai T, Small I (2009) A study of new Arabidopsis chloroplast RNA editing mutants reveals general features of editing factors and their target sites. Plant Cell 21:3686–3699CrossRefPubMedGoogle Scholar
  21. Hayes ML, Reed ML, Hegeman CE, Hanson MR (2006) Sequence elements critical for efficient RNA editing of a tobacco chloroplast transcript in vivo and in vitro. Nucleic Acids Res 34:3742–3754CrossRefPubMedGoogle Scholar
  22. Holec S, Lange H, Kuhn K, Alioua M, Borner T, Gagliardi D (2006) Relaxed transcription in Arabidopsis mitochondria is counterbalanced by RNA stability control mediated by polyadenylation and polynucleotide phosphorylase. Mol Cell Biol 26:2869–2876CrossRefPubMedGoogle Scholar
  23. Hughes AL, Nei M (1989) Nucleotide substitution at major histocompatibility complex class II loci: evidence for overdominant selection. Proc Natl Acad Sci USA 86:958–962CrossRefPubMedGoogle Scholar
  24. Ikemura T (1985) Codon usage and tRNA content in unicellular and multicellular organisms. Mol Biol Evol 2:13–34PubMedGoogle Scholar
  25. Kimchi-Sarfaty C, Oh JM, Kim IW, Sauna ZE, Calcagno AM, Ambudkar SV, Gottesman MM (2007) A “silent” polymorphism in the MDR1 gene changes substrate specificity. Science 315:525–528CrossRefPubMedGoogle Scholar
  26. Kimura M (1983) The neutral theory of molecular evolution. Cambridge University Press, CambridgeCrossRefGoogle Scholar
  27. Kobayashi Y, Matsuo M, Sakamoto K, Wakasugi T, Yamada K, Obokata J (2008) Two RNA editing sites with cis-acting elements of moderate sequence identity are recognized by an identical site-recognition protein in tobacco chloroplasts. Nucleic Acids Res 36:311–318CrossRefPubMedGoogle Scholar
  28. Kotera E, Tasaka M, Shikanai T (2005) A pentatricopeptide repeat protein is essential for RNA editing in chloroplasts. Nature 433:326–330CrossRefPubMedGoogle Scholar
  29. Kubo T, Newton KJ (2008) Angiosperm mitochondrial genomes and mutations. Mitochondrion 8:5–14CrossRefPubMedGoogle Scholar
  30. Kudla G, Murray AW, Tollervey D, Plotkin JB (2009) Coding-sequence determinants of gene expression in Escherichia coli. Science 324:255–258CrossRefPubMedGoogle Scholar
  31. Li WH, Gojobori T, Nei M (1981) Pseudogenes as a paradigm of neutral evolution. Nature 292:237–239CrossRefPubMedGoogle Scholar
  32. Liu Q, Feng Y, Xue Q (2004) Analysis of factors shaping codon usage in the mitochondrion genome of Oryza sativa. Mitochondrion 4:313–320CrossRefPubMedGoogle Scholar
  33. Löhne C, Borsch T (2005) Molecular evolution and phylogenetic utility of the petD group II intron: a case study in basal angiosperms. Mol Biol Evol 22:317–332CrossRefPubMedGoogle Scholar
  34. Lynch M, Koskella B, Schaack S (2006) Mutation pressure and the evolution of organelle genomic architecture. Science 311:1727–1730CrossRefPubMedGoogle Scholar
  35. McDonald JH, Kreitman M (1991) Adaptive protein evolution at the Adh locus in Drosophila. Nature 351:652–654CrossRefPubMedGoogle Scholar
  36. Michel F, Umesono K, Ozeki H (1989) Comparative and functional anatomy of group II catalytic introns—a review. Gene 82:5–30CrossRefPubMedGoogle Scholar
  37. Morton BR (2003) The role of context-dependent mutations in generating compositional and codon usage bias in grass chloroplast DNA. J Mol Evol 56:616–629CrossRefPubMedGoogle Scholar
  38. Morton RA, Morton BR (2007) Separating the effects of mutation and selection in producing DNA skew in bacterial chromosomes. BMC Genomics 8:369CrossRefPubMedGoogle Scholar
  39. Mower JP, Touzet P, Gummow JS, Delph LF, Palmer JD (2007) Extensive variation in synonymous substitution rates in mitochondrial genes of seed plants. BMC Evol Biol 7:135Google Scholar
  40. Mulligan RM, Chang KLC, Chou CC (2007) Computational analysis of RNA editing sites in plant mitochondrial genomes reveals similar information content and a sporadic distribution of editing sites. Mol Biol Evol 24:1971–1981CrossRefPubMedGoogle Scholar
  41. Okuda K, Chateigner-Boutin AL, Nakamura T, Delannoy E, Sugita M, Myouga F, Motohashi R, Shinozaki K, Small I, Shikanai T (2009) Pentatricopeptide repeat proteins with the DYW motif have distinct molecular functions in RNA editing and RNA cleavage in Arabidopsis chloroplasts. Plant Cell 21:146–156CrossRefPubMedGoogle Scholar
  42. Palmer JD, Herbon LA (1988) Plant mitochondrial DNA evolves rapidly in structure, but slowly in sequence. J Mol Evol 28:87–97CrossRefPubMedGoogle Scholar
  43. Parkinson CL, Mower JP, Qiu YL, Shirk AJ, Song K, Young ND, DePamphilis CW, Palmer JD (2005) Multiple major increases and decreases in mitochondrial substitution rates in the plant family Geraniaceae. BMC Evol Biol 5:73Google Scholar
  44. Peden JF (2000) Analysis of codon usage, PhD Thesis. University of NottinghamGoogle Scholar
  45. Petrov DA, Lozovskaya ER, Hartl DL (1996) High intrinsic rate of DNA loss in Drosophila. Nature 384:346–349CrossRefPubMedGoogle Scholar
  46. Picardi E, Regina TM, Brennicke A, Quagliariello C (2007) REDIdb: the RNA editing database. Nucleic Acids Res 35:D173–D177CrossRefPubMedGoogle Scholar
  47. Qiu YL, Cho Y, Cox JC, Palmer JD (1998) The gain of three mitochondrial introns identifies liverworts as the earliest land plants. Nature 394:671–674CrossRefPubMedGoogle Scholar
  48. Ran JH, Gao H, Wang XQ (2010) Fast evolution of the retroprocessed mitochondrial rps3 gene in Conifer II and further evidence for the phylogeny of gymnosperms. Mol Phylogenet Evol 54:136–149CrossRefPubMedGoogle Scholar
  49. Rüdinger M, Polsakiewicz M, Knoop V (2008) Organellar RNA editing and plant-specific extensions of pentatricopeptide repeat proteins in jungermanniid but not in marchantiid liverworts. Mol Biol Evol 25:1405–1414CrossRefPubMedGoogle Scholar
  50. Rüdinger M, Funk HT, Rensing SA, Maier UG, Knoop V (2009) RNA editing: only eleven sites are present in the Physcomitrella patens mitochondrial transcriptome and a universal nomenclature proposal. Mol Genet Genomics 281:473–481CrossRefPubMedGoogle Scholar
  51. Schmid M, Davison TS, Henz SR, Pape UJ, Demar M, Vingron M, Scholkopf B, Weigel D, Lohmann JU (2005) A gene expression map of Arabidopsis thaliana development. Nat Genet 37:501–506CrossRefPubMedGoogle Scholar
  52. Sharp PM, Li WH (1987) The rate of synonymous substitution in enterobacterial genes is inversely related to codon usage bias. Mol Biol Evol 4:222–230PubMedGoogle Scholar
  53. Sharp PM, Tuohy TM, Mosurski KR (1986) Codon usage in yeast: cluster analysis clearly differentiates highly and lowly expressed genes. Nucleic Acids Res 14:5125–5143CrossRefPubMedGoogle Scholar
  54. Shimada H, Sugiura M (1991) Fine structural features of the chloroplast genome: comparison of the sequenced chloroplast genomes. Nucleic Acids Res 19:983–995CrossRefPubMedGoogle Scholar
  55. Sloan DB, Barr CM, Olson MS, Keller SR, Taylor DR (2008) Evolutionary rate variation at multiple levels of biological organization in plant mitochondrial DNA. Mol Biol Evol 25:243–246CrossRefPubMedGoogle Scholar
  56. Sloan DB, Oxelman B, Rautenberg A, Taylor DR (2009) Phylogenetic analysis of mitochondrial substitution rate variation in the angiosperm tribe Sileneae (Caryophyllaceae). BMC Evol Biol 9:260CrossRefPubMedGoogle Scholar
  57. Stajich JE, Block D, Boulez K, Brenner SE, Chervitz SA, Dagdigian C, Fuellen G, Gilbert JG, Korf I, Lapp H, Lehvaslaiho H, Matsalla C, Mungall CJ, Osborne BI, Pocock MR, Schattner P, Senger M, Stein LD, Stupka E, Wilkinson MD, Birney E (2002) The Bioperl toolkit: Perl modules for the life sciences. Genome Res 12:1611–1618CrossRefPubMedGoogle Scholar
  58. Stupar RM, Lilly JW, Town CD, Cheng Z, Kaul S, Buell CR, Jiang J (2001) Complex mtDNA constitutes an approximate 620-kb insertion on Arabidopsis thaliana chromosome 2: implication of potential sequencing errors caused by large-unit repeats. Proc Natl Acad Sci USA 98:5099–5103CrossRefPubMedGoogle Scholar
  59. Takenaka M, Neuwirt J, Brennicke A (2004) Complex cis-elements determine an RNA editing site in pea mitochondria. Nucleic Acids Res 32:4137–4144CrossRefPubMedGoogle Scholar
  60. Tillich M, Lehwark P, Morton BR, Maier UG (2006) The evolution of chloroplast RNA editing. Mol Biol Evol 23:1912–1921CrossRefPubMedGoogle Scholar
  61. Unseld M, Marienfeld JR, Brandt P, Brennicke A (1997) The mitochondrial genome of Arabidopsis thaliana contains 57 genes in 366, 924 nucleotides. Nat Genet 15:57–61CrossRefPubMedGoogle Scholar
  62. Wolfe KH, Li WH, Sharp PM (1987) Rates of nucleotide substitution vary greatly among plant mitochondrial, chloroplast, and nuclear DNAs. Proc Natl Acad Sci USA 84:9054–9058CrossRefPubMedGoogle Scholar
  63. Wright F (1990) The ‘effective number of codons’ used in a gene. Gene 87:23–29CrossRefPubMedGoogle Scholar
  64. Yang Z (2007) PAML 4: phylogenetic analysis by maximum likelihood. Mol Biol Evol 24:1586–1591CrossRefPubMedGoogle Scholar
  65. Yang Z, Nielsen R (2008) Mutation-selection models of codon substitution and their use to estimate selective strengths on codon usage. Mol Biol Evol 25:568–579CrossRefPubMedGoogle Scholar
  66. Yu W, Schuster W (1995) Evidence for a site-specific cytidine deamination reaction involved in C to U RNA editing of plant mitochondria. J Biol Chem 270:18227–18233CrossRefPubMedGoogle Scholar
  67. Zehrmann A, Verbitskiy D, van der Merwe JA, Brennicke A, Takenaka M (2009) A DYW domain-containing pentatricopeptide repeat protein is required for RNA editing at multiple sites in mitochondria of Arabidopsis thaliana. Plant Cell 21:558–567CrossRefPubMedGoogle Scholar
  68. Zhang WJ, Zhou J, Li ZF, Wang L, Gu X, Zhong Y (2007) Comparative analysis of codon usage patterns among mitochondrion, chloroplast and nuclear genes in Triticum aestivum L. J Integr Plant Biol 49:246–254CrossRefGoogle Scholar
  69. Zhou M, Li X (2009) Analysis of synonymous codon usage patterns in different plant mitochondrial genomes. Mol Biol Rep 36:2039–2046CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  1. 1.Department of BiologyUniversity of VirginiaCharlottesvilleUSA

Personalised recommendations