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Molecular and General Genetics MGG

, Volume 246, Issue 3, pp 353–359 | Cite as

A multigene family related to chitin synthase genes of yeast in the opportunistic pathogen Aspergillus fumigatus

  • Emilia Mellado
  • Agnès Aufauvre-Brown
  • Charles A. Specht
  • Phillips W. Robbins
  • David W. Holden
Original Paper

Abstract

Two approaches were used to isolate fragments of chitin synthase genes from the opportunistic human pathogen Aspergillus fumigatus. Firstly, regions of amino acid conservation in chitin synthases of Saccharomyces cerevisiae were used to design degenerate primers for amplification of portions of related genes, and secondly, a segment of the S. cerevisiae CSD2 gene was used to screen an A. fumigatus λ genomic DNA library. The polymerase chain reaction (PCR)-based approach led to the identification of five different genes, designated chsA, chsB, chsC, chsD and chsE. chsA, chsB, and chsC fall into Classes I, II and III of the ‘zymogen type’ chitin synthases, respectively. The chsD fragment has approximately 35% amino acid sequence identity to both the zymogen type genes and the non-zymogen type CSD2 gene. chsF appears to be a homologue of CSD2, being 80% identical to CSD2 over 100 amino acids. An unexpected finding was the isolation by heterologous hybridization of another gene (chsE), which also has strong sequence similarity (54% identity at the amino acid level over the same region as chsF) to CSD2. Reverse transcriptase-PCR was used to show that each gene is expressed during hyphal growth in submerged cultures.

Key words

Chitin synthase Multigene family Fungi 

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References

  1. Archer DB (1977) Chitin biosynthesis in protoplast and subcellular fractions of Aspergillus fumigatus. Biochem J 164:653–658Google Scholar
  2. Au-Young J, Robbins PW (1990) Isolation of a chitin synthase gene (chs1) from Candida albicans by expression in Saccharomyces cerevisiae. Mol Microbiol 4:197–207Google Scholar
  3. Bowen AR, Chen-Wu JL, Momany M, Young R, Szaniszlo PJ, Robbins PW (1992) Classification of fungal chitin synthases. Proc Natl Acad Sci USA 89:519–523Google Scholar
  4. Bulawa CE (1992) CSD2, CSD3, and CSD4, genes required for chitin synthesis in Saccharomyces cerevisiae: the CSD2 gene product is related to chitin synthases and to developmentally regulated proteins in Rhizobium species and Xenopus laevis. Mol Cell Biol 12:1764–1776Google Scholar
  5. Bulawa CE (1993) Genetics and molecular biology of chitin synthesis in fungi. Ann Rev Microbiol 47:505–534Google Scholar
  6. Bulawa CE, Osmond BC (1990) Chitin synthase I and chitin synthase II are not required for chitin synthesis in vivo in Saccharomyces cerevisiae. Proc Natl Acad Sci USA 87:7424–7428Google Scholar
  7. Bulawa CE, Slater M, Cabib E, Au-Young J, Sburlati A, Adair WL, Robbins PW (1986) The S. cerevisiae structural gene for chitin synthase is not required for chitin synthesis in vivo. Cell 46:213–225Google Scholar
  8. Cabib E, Farkas V (1971) The control of morphogenesis: an enzymatic mechanism for the initiation of septum formation in yeast. Proc Natl Acad Sci USA 68:2052–2056Google Scholar
  9. Cabib E, Roberts R, Bowers B (1982) Synthesis of the yeast cell wall and its regulation. Ann Rev Biochem 51:763–793Google Scholar
  10. Chen-Wu JL, Zwicker J, Bowen AR, Robbins PW (1992) Expression of chitin synthase genes during yeast and hyphal growth phases of Candida albicans. Mol Microbiol 6:497–502Google Scholar
  11. Devereux J, Haeberli P, Smithies O (1984) A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12:387–395Google Scholar
  12. Farkas V (1979) Biosynthesis of cell walls of fungi. Microbiol Rev 43:117–144Google Scholar
  13. Fonzi WA, Sypherd PS (1985) Expression of the gene for ornithine decarboxylase of Saccharomyces cerevisiae in Escherichia coli. Mol Cell Biol 5:161–166Google Scholar
  14. Gooday GW (1977) Biosynthesis of the fungal cell wall. Mechanisms and implications. J Gen Microbiol 99:1–11Google Scholar
  15. Gurr SJ, Unkles SE, Kinghorn JR (1987) The structure and organization of nuclear genes of filamentous fungi. In: Kinghorn JR (ed) Gene structure in Eukaryotic Microbes. IRL Press. Oxford, UK, pp. 93–139Google Scholar
  16. Hector RF (1993) Compounds active against cell walls of medically important fungi. Clin Microbiol Rev 6:1–21Google Scholar
  17. Holden DW, Kronstad JW, Leong SA (1989) Mutation in a heat regulated hsp70 gene of Ustilago maydis. EMBO J 8:1927–1934Google Scholar
  18. Higgins DG, Sharp PM (1988) CLUSTAL: a package for performing multiple sequence alignments on a microcomputer. Gene 73:237–244Google Scholar
  19. Katz D, Rosenberger RF (1971) Lysis of an Aspergillus nidulans mutant blocked in chitin synthesis and its relation to wall assembly and wall metabolism. Arch Mikrobiol 80:284–292Google Scholar
  20. Orlean P (1987) Two chitin synthases in Saccharomyces cerevisiae. J Biol Chem 262:5732–5739Google Scholar
  21. Pearson WR (1990) Rapid and sensitive sequence comparison with FASTP and FASTA. Methods Enzymol 183:63–98Google Scholar
  22. Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning. A laboratory manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New YorkGoogle Scholar
  23. Sanger F, Nicklen S, Coulson AR (1977) DNA sequencing with chain terminating inhibitors. Proc Natl Acad Sci USA 74:5463–5467Google Scholar
  24. Shaw JA, Mol PC, Bowers B, Silverman SJ, Valdivieso MH, Durán A, Cabib E (1991) The function of chitin synthases 2 and 3 in the Saccharomyces cerevisiae cell cycle. J Cell Biol 114:111–123Google Scholar
  25. Silverman SJ (1989) Similar and different domains of chitin synthase 1 and 2 of Saccharomyces cerevisiae: two isoenzymes with different functions. Yeast 5:459–467Google Scholar
  26. Silverman SJ, Sburlati A, Slater ML, Cabib E (1988) Chitin synthase 2 is essential for septum formation and cell division in Saccharomyces cerevisiae. Proc Natl Acad Sci USA 85:4735–4739Google Scholar
  27. Sudoh M, Nagahashi S, Doi M, Ohta A, Takagi M, Arisawa M (1993) Cloning of the chitin synthase 3 from Candida albicans and its expression during yeast-hyphal transition. Mol Gen Genet 241:351–358Google Scholar
  28. Tang CM, Cohen J, Holden DW (1992) An Aspergillus fumigatus alkaline protease mutant constructed by gene disruption is deficient in extracellular elastase activity. Mol Microbiol 6:1663–1671Google Scholar
  29. Valdivieso MH, Mol PC, Shaw JA, Cabib E, Durán A (1991) CAL1, A gene required for activity of chitin synthase 3 in Saccharomyces cerevisiae. J Cell Biol 114:101–109Google Scholar
  30. Yarden O, Yanofsky C (1991) Chitin synthase 1 plays a major role in cell wall biogenesis in Neurospora crassa. Genes Dev 5:2420–2430Google Scholar

Copyright information

© Springer-Verlag 1995

Authors and Affiliations

  • Emilia Mellado
    • 1
  • Agnès Aufauvre-Brown
    • 1
  • Charles A. Specht
    • 2
  • Phillips W. Robbins
    • 2
  • David W. Holden
    • 1
  1. 1.Department of Infectious Diseases and Bacteriology, Royal Postgraduate Medical School, Hammersmith HospitalLondonUK
  2. 2.Center for Cancer Research, Massachusetts Institute of TechnologyCambridgeUSA

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