Plant Molecular Biology

, Volume 29, Issue 1, pp 63–80 | Cite as

Two copies of a DNA element, ‘Wendy’, in the chloroplast chromosome of Chlamydomonas reinhardtii between rearranged gene clusters

  • Wen-Hua Fan
  • Mark A. Woelfle
  • Gisela Mosig
Research Article


We have characterized two copies of a 2.4 kb DNA element that we call ‘Wendy’, in the chloroplast chromosome of Chlamydomonas reinhardtii. The two copies of Wendy reside in different single-copy regions at opposite positions in the chloroplast genome. Like many mobile DNA elements, both copies of Wendy are bordered by inverted repeats and contain several additional degenerate copies of these repeat sequences in direct or inverted orientation. In addition, four basepairs are repeated in direct orientation. Two major open reading frames (ORFs) are predicted from the DNA sequence of Wendy I. These ORFs are co-transcribed from a promoter inside the element. The deduced amino acid sequence of the larger of these ORFs shares some weak similarities with sequence motifs of transposases and integrases of other mobile elements. Wendy II appears to be altered relative to Wendy I by point mutations and small deletions and insertions which destroy the ORFs. The leader sequence of the Wendy transcript is nearly identical with the leader sequence of the rbcL transcript of C. reinhardtii, but not of C. moewusii (where the complete Wendy was also undetectable). Furthermore, both copies of Wendy are bracketed by gene clusters that are separated in C. reinhardtii but are contiguous in C. moewusii where they exist in an inverted orientation compared with C. reinhardtii. Wendy was not found in any of the completely sequenced chloroplast genomes of rice, tobacco, pine, Euglena or Marchantia, nor in any other GenBank entry. Our results suggest that Wendy has invaded C. reinhardtii after divergence from other species. Subsequent Wendy-dependent illegitimate homologous or site-specific recombination events or both may have contributed to scrambling of the C. reinhardtii chloroplast genome relative to genomes of other species.

Key words

evolution Chlamydomonas reinhardtii chloroplast site-specific recombination transcription transposition 



open reading frame




large inverted repeat of the chloroplast genome


terminal inverted repeats of Wendy


the TIR near the 5′ end of Wendy


the TIR near the 3′ end of Wendy I


direct repeats bordering Wendy


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    BakerTA: Bacteriophage Mu: a bacteriophage that integrates like a retrovirus. Semin Virol 6: 53–63 (1995).Google Scholar
  2. 2.
    BaldaufSL, PalmerJD: Evolutionary transfer of the chloroplast tufA gene to the nucleus, Nature 344: 262–265 (1990).CrossRefPubMedGoogle Scholar
  3. 3.
    BergD, HoweM (eds): Mobile DNA. American Society for Microbiology, Washington, D.C. (1989).Google Scholar
  4. 4.
    BoudreauE, OtisC, TurmelM: Conserved gene clusters in the highly rearranged chloroplast genomes of Chlamydomonas moewusii and Chlamydomonas reinhardtii. Plant Mol Biol 24: 585–602 (1994).CrossRefPubMedGoogle Scholar
  5. 5.
    ChenE, SeeburgPH: Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA 4: 165–170 (1985).PubMedGoogle Scholar
  6. 6.
    CleggMT, GautBS, LearnGH, MortonBR: Rates and patterns of chloroplast DNA evolution. Proc Natl Acad Sci USA 91: 6795–6801 (1994).PubMedGoogle Scholar
  7. 7.
    CraigN: Transposon Tn7. In: BergD, HoweM (eds) Mobile DNA, pp. 211–225. American Society for Microbiology, Washington, D.C. (1989).Google Scholar
  8. 8.
    DevereuxJ, HaeberliP, SmithiesO: A comprehensive set of sequence analysis programs for the VAX. Nucl Acids Res 12: 387–395 (1984).PubMedGoogle Scholar
  9. 9.
    DronM, RahireM, RochaixJD: Sequence of the chloroplast DNA region of Chlamydomonas reinhardtii containing of gene of the large subunit of ribulose bisphosphate carboxylase and part of its flanking genes. J Mol Biol 162: 775–793 (1982).PubMedGoogle Scholar
  10. 10.
    DürrenbergerF, RochaixJD: Chloroplast ribosomal intron of Chlamydomonas reinhardtii: in vitro self-splicing, DNA endonuclease activity and in vivo mobility. EMBO J 10: 3495–3501 (1991).PubMedGoogle Scholar
  11. 11.
    EvansLR, BrownNL: Construction of hybrid Tn501/Tn21 transposases in vivo: identification of a region of transposase conferring specificity of recognition of the 38 bp terminal inverted repeats. EMBO J 6: 2849–2853 (1987).PubMedGoogle Scholar
  12. 12.
    Fan W-H: A novel transposon-like element in the chloroplast genome of Chlamydomonas reinhardtii. Ph. D. thesis, Vanderbilt University, Nashville, TN (1994).Google Scholar
  13. 13.
    FedoroffN, ShläppiM, RainaR: Epigenetic regulation of the maize Spm transposon. Bioessays 17: 291–297 (1995).PubMedGoogle Scholar
  14. 14.
    FedoroffNV: Maize transposable elements. In: BergD, HoweM (eds) Mobile DNA, pp. 375–411. American Society for Microbiology, Washington, D. C. (1989).Google Scholar
  15. 15.
    GrantDM, GillhamNW, BoyntonJE: Inheritance of chloroplast DNA in Chlamydomonas reinhardtii. Proc Natl Acad Sci USA 77: 6067–6071 (1980).Google Scholar
  16. 16.
    GrayMW: The endosymbiont hypothesis revisited. Int Rev Cytol 141: 233–357 (1992).PubMedGoogle Scholar
  17. 17.
    GroenenMAM, van dePutteP: Analysis of the attachment sites of bacteriophage Mu, using site-directed mutagenesis. J Mol Biol 189: 597–602 (1986).CrossRefPubMedGoogle Scholar
  18. 18.
    HallRM, BrownHJ, BrookesDE, StokesHW: Integrons found in different locations have identical 5′ ends but variable 3′ ends. J Bact 176: 6286–6294 (1994).PubMedGoogle Scholar
  19. 19.
    HallickRB, LongL, DragerRG, FavreauMR, MonfortA, OrsatB, SpielmannA, StutsE: Complete sequence of Euglena gracilis chloroplast DNA. Nucl Acids Res 21: 3537–3544 (1993).PubMedGoogle Scholar
  20. 20.
    HarrisE: Chlamydomonas reinhardtii chloroplast genome. In: O'BrianS J (ed) Genetic Maps, pp. 2.165–2.167. Cold Spring Harbor Press, Cold spring Harbor, NY (1993).Google Scholar
  21. 21.
    HarrisEH: The Chlamydomonas Sourcebook: A Comprehensive Guide to Biology and Laboratory Use. Academic Press, New York (1989).Google Scholar
  22. 22.
    HarrisEH, BoyntonJE, GillhamNW, BurkhartBD: Chloroplast genome organization in Chlamydomonas. Arch Protistenkunde 139: 183–192 (1991).Google Scholar
  23. 23.
    JohnsonCH, SchmidtGW: The psbB gene cluster of Chlamydomonas reinhardtii chloroplast: sequence and transcriptional analyses of psbN and psbH. Plant Mol Biol 22: 645–658 (1993).PubMedGoogle Scholar
  24. 24.
    KhanEJ, MackJPG, KatzPA, KulkoskyJ, SkalkaAM: Retroviral integrase domains: DNA binding and the recognition of LTR sequences. Nucl Acids Res 19: 851–860 (1990).Google Scholar
  25. 24a.
    KhrebtukovaI, SpreitzerRJ: Chlamydomonas chloroplast trnR, trnT, and trnE genes. Plant Physiol 104: 1093–1094 (1994).CrossRefPubMedGoogle Scholar
  26. 25.
    KleinU, SalvadorML, BogoralL: Activity of the Chlamydomonas chloroplast rbcL gene promoter is enhanced by a remote sequence element. Proc Natl Acad Sci USA 91: 10819–10823 (1994).PubMedGoogle Scholar
  27. 26.
    KückU, ChoquetY, SchneiderM, DronM, BennounP: Structural and transcription analysis of two homologous genes for the P700 chlorophyll α-apoproteins in Chlamydomonas reinhardtii: evidence for in vivo trans-splicing. EMBO J 6: 2185–2195 (1987).Google Scholar
  28. 27.
    LambowitzAM, BelfortM: Introns as mobile genetic elements. Annu Rev Biochem 62: 587–622 (1993).CrossRefPubMedGoogle Scholar
  29. 28.
    LavoieBD, ChenBS, AllisonRG, ChaconasG: Structural aspects of a higher order nucleoprotein complex: induction of an altered DNA structure at the Mu-host junction of the Mu type 1 transpososome. EMBO J 10: 3051–3059 (1991).PubMedGoogle Scholar
  30. 29.
    LenichAG, GlasgowAC: Amino acid sequence homology between Piv, an essential protein in site-specific DNA inversion in Moraxella lacunata, and transposases of an unusual family of insertion elements. J Bact 176: 4160–4164 (1994).PubMedGoogle Scholar
  31. 30.
    LipmanDJ, PearsonWR: Rapid and sensitive protein similarity searches. Science 227: 1435–1441 (1985).PubMedGoogle Scholar
  32. 31.
    LoizosN, TillierERM, BelfortM: Evolution of mobile group I intron: Recognition of intron sequences by an intron-encoded endonuclease. Proc Natl Acad Sci USA 91: 11983–11987 (1994).PubMedGoogle Scholar
  33. 32.
    MaasWK: The arginine repressor of Escherichia coli. Microbiol Rev 58: 631–640 (1994).PubMedGoogle Scholar
  34. 33.
    MetsLJ, GeistLJ: Linkage of a known chloroplast gene mutation to the uniparental genome of Chlamydomonas reinhardii. Genetics 105: 559–579 (1983).Google Scholar
  35. 34.
    MetzenbergAB, WurzerG, HuismanTHJ, SmithiesO: Homology requirements for unequal crossing-over in humans. Genetics 128: 143–161 (1991).PubMedGoogle Scholar
  36. 35.
    MilliganBG, HamptonJN, PalmerJD: Dispersed repeats and structural reorganization in subclover chloroplast DNA. Mol Biol Evol 6: 355–368 (1989).PubMedGoogle Scholar
  37. 36.
    MizuuchiM, BakerTA, MizuuchiK: Assembly of the active form of the transposase-Mu DNA complex: A critical control point in Mu transposition. Cell 70: 303–311 (1992).CrossRefPubMedGoogle Scholar
  38. 37.
    NagDK, BergDE: Specificity of bacteriophage Mu excision. Mol Gen Genet 207: 395–401 (1987).CrossRefPubMedGoogle Scholar
  39. 38.
    O'NeillGP, SchonA, ChowA, ChenMW, KimY, SöllD: Sequence of tRNAGlu and its genes from the chloroplast genome of Chlamydomonas reinhardtii.. Nucl Acids Res 18: 5893 (1990).PubMedGoogle Scholar
  40. 39.
    PalmerJD: Plastid chromosomes: structure and evolution. In: VasilK, BogoradL (eds) The Molecular Biology of Plastids, pp. 5–53. Academic Press, San Diego (1991).Google Scholar
  41. 40.
    PalmerJD, BoyntonJE, GillhamNW, HarrisEH: Evolution and recombination of the large inverted repeat in Chlamydomonas chloroplast DNA. In: SteinbackKE, BonitzS, ArntzenCJ, BogoradL (eds) Molecular Biology of the Photosynthetic Apparatus pp. 269–278. Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1985).Google Scholar
  42. 41.
    PatoML: Bacteriophage Mu. In: BergD, HoweM (eds) Mobile DNA, pp. 23–52. American Society for Microbiology, Washington, DC (1989).Google Scholar
  43. 42.
    PlasterkRH: Frameshift control of IS1 transposition. Trends Genet 7: 203–206 (1991).PubMedGoogle Scholar
  44. 43.
    PleijCWA: Pseudoknots: a new motif in the RNA game. Trends biochem 15: 143–147 (1990).CrossRefGoogle Scholar
  45. 44.
    RadmanM: Avoidance of inter-repeat recombination by sequence divergence and a mechanism of neutral evolution. Biochimie 73: 357–361 (1991).CrossRefPubMedGoogle Scholar
  46. 45.
    ReznikoffWS, McClureWR: E. coli promoters. In: ReznikoffW, GoldL (eds) Maximizing Gene Expression, pp. 1–33. Butterworth, Seven Oaks, UK (1986).Google Scholar
  47. 46.
    RioDC, LaskiFA, RubinGM: Identification and immunochemical analysis of biologically active Drosophila P element transposase. Cell 44: 21–32 (1986).CrossRefPubMedGoogle Scholar
  48. 47.
    RochaixJD, RahireM, MichelF: The chloroplast ribosomal intron of Chlamydomonas reinhardtii codes for a polypeptide related to mitochondrial maturase. Nucl Acids Res 13: 975–984 (1985).PubMedGoogle Scholar
  49. 48.
    SambrookJ, FritschEF, ManiatisT: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Press, Cold Spring Harbor, NY (1989).Google Scholar
  50. 49.
    SchneiderM, RochaixJD: Sequence organization of the chloroplast ribosomal spacer of Chlamydomonas reinhardtii: uninterrupted tRNAI1e and tRNA AA1a genes and extensive secondary structure. Plant Mol Biol 6: 265–270 (1986).Google Scholar
  51. 50.
    ScottJR, BringelF, RudyCK: Conjugative trasposition of Tn916: preferred targets and evidence for conjugative transfer of a single strand and for a double-stranded circular intermediate. Mol Microbiol 11: 1099–1108 (1994).PubMedGoogle Scholar
  52. 51.
    SelkerEU: Premeiotic instability of repeated sequences in Neurospora crassa. Annu Rev Genet 24: 579–613 (1990).CrossRefPubMedGoogle Scholar
  53. 52.
    ShapiroJA, LeachD: Action of a transposable element in coding sequence fusions. Genetics 126: 293–299 (1990).PubMedGoogle Scholar
  54. 53.
    SternDB, KindleK: 3′ End maturation of the Chlamydomonas reinhardtii chloroplast atpB mRNA is a two step process. Mol Cell Biol 13: 2277–2285 (1993).PubMedGoogle Scholar
  55. 54.
    StirlingCJ, SzatmariG, StewartG, SmithMCM, SherrattDJ: The arginine repressor is essential for plasmid-stabilizing site-specific recombination at the ColE1 cer locus. EMBO J 7: 4389–4395 (1988).PubMedGoogle Scholar
  56. 55.
    StokesHW, HallRM: A novel family of potentially mobile DNA elements encoding site-specific gene-integration functions: integrons. Mol Microbiol 3: 1669–1683 (1989).PubMedGoogle Scholar
  57. 56.
    SymondsN, ToussaintA, van dePutteP, HoweM (eds): Phage Mu. Cold Spring Harbor Laboratory, Cold spring Harbor, NY (1987).Google Scholar
  58. 57.
    ThompsonJF, LandyA: Regulation of bacteriophage lambda site-specific recombination. In: BergD, HoweM (eds) Mobile DNA, pp. 1–22. American Society for Microbiology, Washington, DC (1989).Google Scholar
  59. 58.
    ThompsonRJ, MosigG: Light and genetic determinants in the control of specific chloroplast transcripts in Chlamydomonas reinhardtii. Plant Physiol 76: 1–6 (1984).Google Scholar
  60. 59.
    ThompsonRJ, MosigG: An ATP-dependent supercoling topoisomerase of Chlamydomonas reinhardtii affects accumulation of specific chloroplast transcripts. Nucl Acids Res 13: 873–891 (1985).PubMedGoogle Scholar
  61. 60.
    TurnerAK, GrinstedJ: DNA sequence of the transposase gene of the new category of class II transposon, Tn2501. Nucl Acids Res 15: 10049 (1987).PubMedGoogle Scholar
  62. 61.
    WakasugiT, TsudzukiJ, ItoS, NakashimaK, TsudzukiT, SugiuraM: Loss of all ndh genes as determined by sequencing the entire chloroplast genome of the black pine Pinus thunbergii. Proc Natl Acad Sci USA 91: 9794–9798 (1994).PubMedGoogle Scholar
  63. 62.
    WeisbergRA, LandyA: Site-specific recombination in phage lambda. In: HendrixRW, RobertsJW, StahlFW, WeisbergRA (eds) Lambda II, pp. 211–250. Cold Spring Harbor Press, Cold Spring Harbor, NY (1983).Google Scholar
  64. 63.
    WernerMH, CloreGM, GronenbornAM, NashHA: Symmetry and asymmetry in the function of Escherichia coli integration host factor: implications for target identification by DNA-binding proteins. Burr Biol 4: 477–487 (1994).Google Scholar
  65. 64.
    WoelfleMA, ThompsonRJ, MosigG: Roles of novobiocin-sensitive topoisomerases in chloroplast DNA replication in Chlamydomonas reinhardtii. Nucl Acids Res 21: 4231–4238 (1993).PubMedGoogle Scholar
  66. 65.
    YangRC, DoveM, SeligyVL, LemieuxC, TurmelM, NarangSA: C. moewusii chloroplast LS gene encoding ribulose-1,5-bisphosphate carboxylase large subunit, and translated products. Gene 50: 259–270 (1986).CrossRefPubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 1995

Authors and Affiliations

  • Wen-Hua Fan
    • 1
  • Mark A. Woelfle
    • 1
  • Gisela Mosig
    • 1
  1. 1.Department of Molecular BiologyVanderbilt UniversityNashvilleUSA

Personalised recommendations