1887

Abstract

The anaerobic gut fungi occupy a unique niche in the intestinal tract of large herbivorous animals and are thought to act as primary colonizers of plant material during digestion. They are the only known obligately anaerobic fungi but molecular analysis of this group has been hampered by difficulties in their culture and manipulation, and by their extremely high A+T nucleotide content. This study begins to answer some of the fundamental questions about the structure and organization of the anaerobic gut fungal genome. Directed plasmid libraries using genomic DNA digested with highly or moderately rich AT-specific restriction enzymes (I and RI) were prepared from a polycentric isolate. Clones were sequenced from these libraries and the breadth of genomic inserts, both genic and intergenic, was characterized. Genes encoding numerous functions not previously characterized for these fungi were identified, including cytoskeletal, secretory pathway and transporter genes. A peptidase gene with no introns and having sequence similarity to a gene encoding a bacterial peptidase was also identified, extending the range of metabolic enzymes resulting from apparent trans-kingdom transfer from bacteria to fungi, as previously characterized largely for genes encoding plant-degrading enzymes. This paper presents the first thorough analysis of the genic, intergenic and rDNA regions of a variety of genomic segments from an anaerobic gut fungus and provides observations on rules governing intron boundaries, the codon biases observed with different types of genes, and the sequence of only the second anaerobic gut fungal promoter reported. Large numbers of retrotransposon sequences of different types were found and the authors speculate on the possible consequences of any such transposon activity in the genome. The coding sequences identified included several orphan gene sequences, including one with regions strongly suggestive of structural proteins such as collagens and lampirin. This gene was present as a single copy in , was expressed during vegetative growth and was also detected in genomes from another gut fungal genus, .

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2005-01-01
2024-12-03
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References

  1. Akhmanova A., Voncken F. G., Harhangi H., Hosea K. M., Vogels G. D., Hackstein J. H. 1998; Cytosolic enzymes with a mitochondrial ancestry from the anaerobic chytrid Piromyces sp. E2. Mol Microbiol 30:1017–1027 [CrossRef]
    [Google Scholar]
  2. Akhmanova A., Voncken F. G., Hosea K. M., Harhangi H., Keltjens J. T., Vogels G. D., Hackstein J. H, Op den Camp H. J. 1999; A hydrogenosome with pyruvate formate-lyase, anaerobic chytrid fungi use an alternative route for pyruvate catabolism. Mol Microbiol 32:1103–1114 [CrossRef]
    [Google Scholar]
  3. Andersson J. O., Roger A. J. 2002; A cyanobacterial gene in nonphotosynthetic protists – an early chloroplast acquisition in eukaryotes?. Curr Biol 12:115–119
    [Google Scholar]
  4. Bayer E. A., Shimon L. J. W., Shoham Y., Lamed R. 1998; Cellulosomes – structure and ultrastructure. J Struct Biol 124:221–234 [CrossRef]
    [Google Scholar]
  5. Billon-Grand G., Fiol J. B., Breton A., Bruyere A., Oulhaj Z. 1991; DNA of some anaerobic rumen fungi, G+C content determination. FEMS Microbiol Lett 66:267–270
    [Google Scholar]
  6. Black G. W., Hazlewood G. P., Xue G. P., Orpin C. G., Gilbert H. J. 1994; Xylanase B from Neocallimastix patriciarum contains a non-catalytic 455-residue linker sequence comprised of 57 repeats of an octapeptide. Biochem J 299:381–387
    [Google Scholar]
  7. Bochicchio B., Pepe A., Tamburro A. M. 2001; On (GGLGY) synthetic repeating sequences of lamprin and analogous sequences. Matrix Biol 20:243–250 [CrossRef]
    [Google Scholar]
  8. Bon E., Casaregola S., Blandin G. 8 other authors 2003; Molecular evolution of eukaryotic genomes: hemiascomycetous yeast spliceosomal introns. Nucleic Acids Res 31:1121–1135 [CrossRef]
    [Google Scholar]
  9. Brookman J. L., Mennim G., Trinci A. P., Theodorou M. K., Tuckwell D. S. 2000; Identification and characterization of anaerobic gut fungi using molecular methodologies based on ribosomal ITS1 and 18S rRNA. Microbiology 146:393–403
    [Google Scholar]
  10. Brownlee A. G. 1989; Remarkably AT-rich genomic DNA from the anaerobic fungus Neocallimastix . Nucleic Acids Res 17:1327–1335 [CrossRef]
    [Google Scholar]
  11. Bulmer M. 1988; Codon usage and intragenic position. J Theor Biol 133:67–71 [CrossRef]
    [Google Scholar]
  12. Celerin M. J. M., Ray J. M., Schisler N. J., Day A. W., Stetler-Stevenson W. G., Laudenbach D. E. 1996; Fungal fimbriae are composed of collagen. EMBO J 15:4445–4453
    [Google Scholar]
  13. Deutsch M., Long M. 1999; Intron-exon structure of eukaryotic model organisms. Nucleic Acids Res 27:3219–3228 [CrossRef]
    [Google Scholar]
  14. Diaz-Lazcoz Y., Henaut A., Vigier P., Risler J. L. 1995; Differential codon usage for conserved amino acids, evidence that the serine codons TCN were primordial. J Mol Biol 250:123–127 [CrossRef]
    [Google Scholar]
  15. Durand R., Fischer M., Rascle C., Fevre M. 1995; Neocallimastix frontalis enolase gene, enol – first report of an intron in an anaerobic fungus. Microbiology 141:1301–1308 [CrossRef]
    [Google Scholar]
  16. Embley T. M., Horner D. S., Dyal P. L., Foster P, van der Giezen M. 2003; Mitochondria and hydrogenosomes are two forms of the same fundamental organelle. Philos Trans Roy Soc Lond B Biol Sci 358:191–203 [CrossRef]
    [Google Scholar]
  17. Farman M. L., Tosa Y., Nitta N., Leong S. A. 1996; MAGGY, a retrotransposon in the genome of the rice blast fungus Magnaporthe grisea . Mol Gen Genet 251:665–674
    [Google Scholar]
  18. Fickett J. W., Tung C. S. 1992; Assessment of protein coding measures. Nucleic Acids Res 20:6441–6450 [CrossRef]
    [Google Scholar]
  19. Fischer M., Durand R., Fevre M. 1995; Characterization of the promoter region of the enolase encoding gene enol from the anaerobic fungus Neocallimastix frontalis – sequence and promoter analysis. Curr Genet 28:80–86 [CrossRef]
    [Google Scholar]
  20. Fontes C. M., Hazlewood G. P., Morag E., Hall J., Hirst B. H., Gilbert H. J. 1995; Evidence for a general role for non-catalytic thermostabilizing domains in xylanases from thermophilic bacteria. Biochem J 307:151–158
    [Google Scholar]
  21. Garcia-Vallvé S., Romeu A., Palau J. 2000; Horizontal gene transfer of glycosyl hydrolases of the rumen fungi. Mol Biol Evol 17:352–361 [CrossRef]
    [Google Scholar]
  22. Gardner M. J. 2001; A status report on the sequencing and annotation of the P. falciparum genome. Mol Biochem Parasitol 118:133–138 [CrossRef]
    [Google Scholar]
  23. Gardner M. J., Hall N., Fing E. 42 other authors 2002a; Genome sequence of the human malaria parasite Plasmodium falciparum . Nature 419:498–511 [CrossRef]
    [Google Scholar]
  24. Gardner M. J., Shallom S. J., Carlton J. M. 34 other authors 2002b; Sequence of Plasmodium falciparum chromosomes 2, 10, 11 and 14. Nature 419:531–534 [CrossRef]
    [Google Scholar]
  25. Grocock R. J., Sharp P. M. 2002; Synonymous codon usage in Pseudomonas aeruginosa PA01. Gene 289:131–139 [CrossRef]
    [Google Scholar]
  26. Hall N., Pain A., Berriman M. 77 other authors 2002; Sequence of Plasmodium falciparum chromosomes 1, 3–9 and 13. Nature 419:527–531 [CrossRef]
    [Google Scholar]
  27. Harhangi H. R., Akhmanova A., Steenbakkers P. J. M., Jetten M. S. M., van der Drift C., Op den Camp H. J. M. 2003a; Genomic DNA analysis of genes encoding (hemi-)cellulolytic enzymes of the anaerobic fungus Piromyces sp. E2. Gene 314:73–80 [CrossRef]
    [Google Scholar]
  28. Harhangi H. R., Freelove A. C., Ubhayasekera W. 8 other authors 2003b; Cel6A, a major exoglucanase from the cellulosome of the anaerobic fungi Piromyces sp. E2 and Piromyces equi . Biochim Biophys Acta 1628:30–39 [CrossRef]
    [Google Scholar]
  29. Harhangi H. R., Akhmanova A., Emmens R. 6 other authors 2003c; Xylose metabolism in the anaerobic fungus Piromyces sp. strain E2 follows the bacterial pathway. Arch Microbiol 180:134–141 [CrossRef]
    [Google Scholar]
  30. Henrich B., Monnerjahn U., Plapp R. 1990; Peptidase D gene ( pepD ) of Escherichia coli K-12, nucleotide sequence, transcript mapping, and comparison with other peptidase genes. J Bacteriol 172:4641–4651
    [Google Scholar]
  31. Kielty C. M., Hopkinson I., Grant M. E. 1993; Collagen. In Connective Tissue and its Heritable Disorders pp 103–147 Edited by Royce P. M., Steinmann B. New York: Wiley-Liss;
    [Google Scholar]
  32. Ko K. S., Jung H. S. 2002; Three nonorthologous ITS1 types are present in a polypore fungus Trichaptum abietinum . Mol Phylogenet Evol 23:112–122 [CrossRef]
    [Google Scholar]
  33. Kocherginskaya S. A., Aminov R. I., White B. A. 2001; Analysis of the rumen bacterial diversity under two different diet conditions using denaturing gradient gel electrophoresis, random sequencing, and statistical ecology approaches. Anaerobe 7:119–134 [CrossRef]
    [Google Scholar]
  34. Lowe S. E., Theodorou M. K., Trinci A. P. J., Hespell R. B. 1985; Growth of anaerobic rumen fungi on defined and semi-defined media lacking rumen fluid. J Gen Microbiol 131:2225–2229
    [Google Scholar]
  35. Moriyama E. N., Powell J. R. 1998; Gene length and codon usage bias in Drosophila melanogaster , Saccharomyces cerevisiae and Escherichia coli . Nucleic Acids Res 26:3188–3193 [CrossRef]
    [Google Scholar]
  36. Munn E. A., Orpin C. G., Greenwood C. A. 1988; The ultrastructure and possible relationships of four obligate anaerobic chytridiomycete fungi from the rumen of sheep. Biosystems 22:67–81 [CrossRef]
    [Google Scholar]
  37. Nakayashiki H., Nishimoto N., Ikeda K., Tosa Y., Mayama S. 1999; Degenerate MAGGY elements in a subgroup of Pyricularia grisea , a possible example of successful capture of a genetic invader by a fungal genome. Mol Gen Genet 261:958–966 [CrossRef]
    [Google Scholar]
  38. Nicholson M. J. 2003; Molecular characterisation of anaerobic gut fungi and their colonisation of plant material in the rumen . PhD thesis Faculty of Science & Engineering, University of Manchester; UK:
  39. Nixon J. E., Wang A., Morrison H. G., McArthur A. G., Sogin M. L., Loftus B. J., Samuelson J. 2002; A spliceosomal intron in Giardia lamblia . Proc Natl Acad Sci U S A 99:3701–3705 [CrossRef]
    [Google Scholar]
  40. O'Donnell K., Cigelnik E. 1997; Two divergent intragenomic rDNA ITS2 types within a monophyletic lineage of the fungus Fusarium are nonorthologous. Mol Phylog Evol 7:103–116 [CrossRef]
    [Google Scholar]
  41. Orpin C. G. 1975; Studies on the rumen flagellate Neocallimastix frontalis . J Gen Microbiol 98:423–430
    [Google Scholar]
  42. Ozkose E., Thomas B. J., Davies D. R., Griffith G. W., Theodorou M. K. 2001; Cyllamyces aberensis gen. nov. sp. nov., a new anaerobic gut fungus with branched sporangiophores isolated from cattle. Can J Bot 79:666–673
    [Google Scholar]
  43. Poulter R., Butler M. 1998; A retrotransposon family from the pufferfish (fugu) Fugu rubripes . Gene 215:241–249 [CrossRef]
    [Google Scholar]
  44. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual , 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  45. Steenbakkers P. J. M., Li X.-L., Ximenes E. A., Arts J. G., Chen H., Ljungdahl L. G., Op den Camp H. J. M. 2001; Noncatalytic docking domains of cellulosomes of anaerobic fungi. J Bacteriol 183:5325–5333 [CrossRef]
    [Google Scholar]
  46. Steenbakkers P. J., Ubhayasekera W., Goossen H. J., Vogels G. D., Mowbray S. L, van Lierop E. M., van der Drift C., Op den Camp H. J. 2002; An intron-containing glycoside hydrolase family 9 cellulase gene encodes the dominant 90 kDa component of the cellulosome of the anaerobic fungus Piromyces sp. strain E2. Biochem J 365:193–204 [CrossRef]
    [Google Scholar]
  47. Theodorou M. K., Mennim G., Davies D., Zhu W.-Y., Trinci A. P. J., Brookman J. 1996; Anaerobic fungi in the digestive tract of mammalian herbivores and their potential for exploitation. Proc Nutrition Society 55:913–926 [CrossRef]
    [Google Scholar]
  48. van der Giezen M., Rechinger K. B., Svendson I., Durand R., Hirt R. P., Prins R. A, Fèvre M., Embley T. M. 1997; A mitochondrial-like targeting signal on the hydrogenosomal malic enzyme from the anaerobic fungus Neocallimastix frontalis : support for the hypothesis that hydrogenosomes are modified mitochondria. Mol Microbiol 23:11–21 [CrossRef]
    [Google Scholar]
  49. van Nues R. W., Venema J., Rientjes J. M., Dirks-Mulder A., Raué H. A. 1995; Processing of eukaryotic pre-rRNA: the role of the transcribed spacers. Biochem Cell Biol 73:789–801 [CrossRef]
    [Google Scholar]
  50. Vinogradov A. E. 2001; Intron length and codon usage. J Mol Evol 52:2–5 [CrossRef]
    [Google Scholar]
  51. Voncken F. G. J., Boxma B., Akhmanova A. S., Vogels G. D., Huynene M., Veenhuis M., Hackstein J. H. P, van Hoek A. H. A. M. 2002; A hydrogenosomal [Fe]-hydrogenase from the anaerobic chytrid Neocallimastix sp. L2. Gene 284:103–112 [CrossRef]
    [Google Scholar]
  52. Winckler T., Dingermann T., Glöckner G. 2002; Dictyostelium mobile elements – strategies to amplify in a compact genome. Cell Mol Life Sci 59:2097–2111 [CrossRef]
    [Google Scholar]
  53. Zhou L., Xue G. P., Orpin C. G., Black G. W., Gilbert H. J., Hazlewood G. P. 1994; Intronless celB from the anaerobic fungus Neocallimastix patriciarum encodes a modular family A endoglucanase. Biochem J 297:359–364
    [Google Scholar]
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