Skip to main content
NCBI home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1996 Dec;62(12):4568–4575. doi: 10.1128/aem.62.12.4568-4575.1996

Characterization of the function of the ver-1A and ver-1B genes, involved in aflatoxin biosynthesis in Aspergillus parasiticus.

S H Liang 1, C D Skory 1, J E Linz 1
PMCID: PMC168283  PMID: 8953728

Abstract

The ver-1A gene was cloned and its nucleotide sequence was determined as part of a previous study on aflatoxin B1 (AFB1) biosynthesis in the filamentous fungus Aspergillus parasiticus SU-1. A second copy of this gene, ver-1B, was tentatively identified in this fungal strain. In this study, ver-1B was cloned by screening an A. parasiticus cosmid library with a ver-1A probe. The nucleotide sequence of ver-1B was determined. The predicted amino acid sequence of ver-1B had 95% identity with ver-1A. A translational stop codon, found in the ver-1B gene coding region, indicated that it encodes a truncated polypeptide. To confirm the function of the ver-1 genes in AFB1 synthesis, a plasmid (pDV-VA) was designed to disrupt ver-1A and/or ver-1B by transformation of the AFB1 producer A. parasiticus NR-1. One disruptant, VAD-102, which accumulated the pathway intermediate versicolorin A was obtained. Southern hybridization analysis of VAD-102 revealed that ver-1A but not ver-1B was disrupted. A functional ver-1A gene was transformed back into strain VAD-102. Transformants which received ver-1A produced AFB1, confirming that ver-1A is the only functional ver-1 gene in A. parasiticus SU-1 and that its gene product is involved in the conversion of versicolorin A to sterigmatocystin in AFB1 biosynthesis. A duplicated chromosomal region (approximately 12 kb) was identified upstream from ver-1A and ver-1B by Southern hybridization analysis. This duplicated region contained the aflR gene, which is proposed to be one regulator of AFB1, synthesis. A similar gene duplication was also identified in several other strains of A. parasiticus.

Full Text

The Full Text of this article is available as a PDF (519.1 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Adamson R. H., Correa P., Sieber S. M., McIntire K. R., Dalgard D. W. Carcinogenicity of aflatoxin B1 in rhesus monkeys: two additional cases of primary liver cancer. J Natl Cancer Inst. 1976 Jul;57(1):67–78. doi: 10.1093/jnci/57.1.67. [DOI] [PubMed] [Google Scholar]
  2. Bennett J. W. Aflatoxins and anthraquinones from diploids of Aspergillus parasiticus. J Gen Microbiol. 1979 Jul;113(1):127–136. doi: 10.1099/00221287-113-1-127. [DOI] [PubMed] [Google Scholar]
  3. Cary J. W., Wright M., Bhatnagar D., Lee R., Chu F. S. Molecular characterization of an Aspergillus parasiticus dehydrogenase gene, norA, located on the aflatoxin biosynthesis gene cluster. Appl Environ Microbiol. 1996 Feb;62(2):360–366. doi: 10.1128/aem.62.2.360-366.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Chang P. K., Cary J. W., Bhatnagar D., Cleveland T. E., Bennett J. W., Linz J. E., Woloshuk C. P., Payne G. A. Cloning of the Aspergillus parasiticus apa-2 gene associated with the regulation of aflatoxin biosynthesis. Appl Environ Microbiol. 1993 Oct;59(10):3273–3279. doi: 10.1128/aem.59.10.3273-3279.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chang P. K., Cary J. W., Yu J., Bhatnagar D., Cleveland T. E. The Aspergillus parasiticus polyketide synthase gene pksA, a homolog of Aspergillus nidulans wA, is required for aflatoxin B1 biosynthesis. Mol Gen Genet. 1995 Aug 21;248(3):270–277. doi: 10.1007/BF02191593. [DOI] [PubMed] [Google Scholar]
  6. Chang P. K., Skory C. D., Linz J. E. Cloning of a gene associated with aflatoxin B1 biosynthesis in Aspergillus parasiticus. Curr Genet. 1992 Mar;21(3):231–233. doi: 10.1007/BF00336846. [DOI] [PubMed] [Google Scholar]
  7. Davis N. D., Iyer S. K., Diener U. L. Improved method of screening for aflatoxin with a coconut agar medium. Appl Environ Microbiol. 1987 Jul;53(7):1593–1595. doi: 10.1128/aem.53.7.1593-1595.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dutton M. F. Enzymes and aflatoxin biosynthesis. Microbiol Rev. 1988 Jun;52(2):274–295. doi: 10.1128/mr.52.2.274-295.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hallam S. E., Malpartida F., Hopwood D. A. Nucleotide sequence, transcription and deduced function of a gene involved in polyketide antibiotic synthesis in Streptomyces coelicolor. Gene. 1988 Dec 30;74(2):305–320. doi: 10.1016/0378-1119(88)90165-5. [DOI] [PubMed] [Google Scholar]
  10. Ichinose K., Kiyono J., Ebizuka Y., Sankawa U. Post-aromatic deoxygenation in polyketide biosynthesis: reduction of aromatic rings in the biosyntheses of fungal melanin and anthraquinone. Chem Pharm Bull (Tokyo) 1993 Nov;41(11):2015–2021. doi: 10.1248/cpb.41.2015. [DOI] [PubMed] [Google Scholar]
  11. Jelinek C. F., Pohland A. E., Wood G. E. Worldwide occurrence of mycotoxins in foods and feeds--an update. J Assoc Off Anal Chem. 1989 Mar-Apr;72(2):223–230. [PubMed] [Google Scholar]
  12. Keller N. P., Kantz N. J., Adams T. H. Aspergillus nidulans verA is required for production of the mycotoxin sterigmatocystin. Appl Environ Microbiol. 1994 May;60(5):1444–1450. doi: 10.1128/aem.60.5.1444-1450.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Keller N. P., Segner S., Bhatnagar D., Adams T. H. stcS, a putative P-450 monooxygenase, is required for the conversion of versicolorin A to sterigmatocystin in Aspergillus nidulans. Appl Environ Microbiol. 1995 Oct;61(10):3628–3632. doi: 10.1128/aem.61.10.3628-3632.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lee L. S., Bennett J. W., Cucullu A. F., Stanley J. B. Synthesis of versicolorin A by a mutant strain of Aspergillus parasiticus deficient in aflatoxin production. J Agric Food Chem. 1975 Nov-Dec;23(6):1132–1134. doi: 10.1021/jf60202a011. [DOI] [PubMed] [Google Scholar]
  15. Mahanti N., Bhatnagar D., Cary J. W., Joubran J., Linz J. E. Structure and function of fas-1A, a gene encoding a putative fatty acid synthetase directly involved in aflatoxin biosynthesis in Aspergillus parasiticus. Appl Environ Microbiol. 1996 Jan;62(1):191–195. doi: 10.1128/aem.62.1.191-195.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Oakley B. R., Rinehart J. E., Mitchell B. L., Oakley C. E., Carmona C., Gray G. L., May G. S. Cloning, mapping and molecular analysis of the pyrG (orotidine-5'-phosphate decarboxylase) gene of Aspergillus nidulans. Gene. 1987;61(3):385–399. doi: 10.1016/0378-1119(87)90201-0. [DOI] [PubMed] [Google Scholar]
  17. Payne G. A., Nystrom G. J., Bhatnagar D., Cleveland T. E., Woloshuk C. P. Cloning of the afl-2 gene involved in aflatoxin biosynthesis from Aspergillus flavus. Appl Environ Microbiol. 1993 Jan;59(1):156–162. doi: 10.1128/aem.59.1.156-162.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Skory C. D., Chang P. K., Cary J., Linz J. E. Isolation and characterization of a gene from Aspergillus parasiticus associated with the conversion of versicolorin A to sterigmatocystin in aflatoxin biosynthesis. Appl Environ Microbiol. 1992 Nov;58(11):3527–3537. doi: 10.1128/aem.58.11.3527-3537.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Skory C. D., Horng J. S., Pestka J. J., Linz J. E. Transformation of Aspergillus parasiticus with a homologous gene (pyrG) involved in pyrimidine biosynthesis. Appl Environ Microbiol. 1990 Nov;56(11):3315–3320. doi: 10.1128/aem.56.11.3315-3320.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Tatebayashi K., Kato J., Ikeda H. Structural analyses of DNA fragments integrated by illegitimate recombination in Schizosaccharomyces pombe. Mol Gen Genet. 1994 Jul 25;244(2):111–119. doi: 10.1007/BF00283511. [DOI] [PubMed] [Google Scholar]
  22. Trail F., Mahanti N., Rarick M., Mehigh R., Liang S. H., Zhou R., Linz J. E. Physical and transcriptional map of an aflatoxin gene cluster in Aspergillus parasiticus and functional disruption of a gene involved early in the aflatoxin pathway. Appl Environ Microbiol. 1995 Jul;61(7):2665–2673. doi: 10.1128/aem.61.7.2665-2673.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Vidal-Cros A., Viviani F., Labesse G., Boccara M., Gaudry M. Polyhydroxynaphthalene reductase involved in melanin biosynthesis in Magnaporthe grisea. Purification, cDNA cloning and sequencing. Eur J Biochem. 1994 Feb 1;219(3):985–992. doi: 10.1111/j.1432-1033.1994.tb18581.x. [DOI] [PubMed] [Google Scholar]
  24. Yu J., Cary J. W., Bhatnagar D., Cleveland T. E., Keller N. P., Chu F. S. Cloning and characterization of a cDNA from Aspergillus parasiticus encoding an O-methyltransferase involved in aflatoxin biosynthesis. Appl Environ Microbiol. 1993 Nov;59(11):3564–3571. doi: 10.1128/aem.59.11.3564-3571.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Yu J., Chang P. K., Cary J. W., Wright M., Bhatnagar D., Cleveland T. E., Payne G. A., Linz J. E. Comparative mapping of aflatoxin pathway gene clusters in Aspergillus parasiticus and Aspergillus flavus. Appl Environ Microbiol. 1995 Jun;61(6):2365–2371. doi: 10.1128/aem.61.6.2365-2371.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES