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. 1997 Aug 15;16(16):4916–4923. doi: 10.1093/emboj/16.16.4916

Aspergillus sporulation and mycotoxin production both require inactivation of the FadA G alpha protein-dependent signaling pathway.

J K Hicks 1, J H Yu 1, N P Keller 1, T H Adams 1
PMCID: PMC1170127  PMID: 9305634

Abstract

The filamentous fungus Aspergillus nidulans contains a cluster of 25 genes that encode enzymes required to synthesize a toxic and carcinogenic secondary metabolite called sterigmatocystin (ST), a precursor of the better known fungal toxin aflatoxin (AF). One ST Cluster (stc) gene, aflR, functions as a pathway-specific transcriptional regulator for activation of other genes in the ST pathway. However, the mechanisms controlling activation of aflR and synthesis of ST and AF are not understood. Here we show that one important level for control of stc gene expression requires genes that were first identified as early acting regulators of asexual sporulation. Specifically, we found that loss-of-function mutations in flbA, which encodes a RGS domain protein, or dominant activating mutations in fadA, which encodes the alpha subunit of a heterotrimeric G protein, block both ST production and asexual sporulation. Moreover, overexpression of flbA or dominant interfering fadA mutations cause precocious stc gene expression and ST accumulation, as well as unscheduled sporulation. The requirement for flbA in sporulation and ST production could be suppressed by loss-of-function fadA mutations. The ability of flbA to activate stc gene expression was dependent upon another early acting developmental regulator, fluG, and AflR, the stc gene-specific transcription factor. These results are consistent with a model in which both asexual sporulation and ST production require inactivation of proliferative growth through inhibition of FadA-dependent signaling. This regulatory mechanism is conserved in AF-producing fungi and could therefore provide a means of controlling AF contamination.

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Selected References

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  1. Adams T. H., Boylan M. T., Timberlake W. E. brlA is necessary and sufficient to direct conidiophore development in Aspergillus nidulans. Cell. 1988 Jul 29;54(3):353–362. doi: 10.1016/0092-8674(88)90198-5. [DOI] [PubMed] [Google Scholar]
  2. Adams T. H., Timberlake W. E. Developmental repression of growth and gene expression in Aspergillus. Proc Natl Acad Sci U S A. 1990 Jul;87(14):5405–5409. doi: 10.1073/pnas.87.14.5405. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Beppu T. Secondary metabolites as chemical signals for cellular differentiation. Gene. 1992 Jun 15;115(1-2):159–165. doi: 10.1016/0378-1119(92)90554-3. [DOI] [PubMed] [Google Scholar]
  4. Berman D. M., Wilkie T. M., Gilman A. G. GAIP and RGS4 are GTPase-activating proteins for the Gi subfamily of G protein alpha subunits. Cell. 1996 Aug 9;86(3):445–452. doi: 10.1016/s0092-8674(00)80117-8. [DOI] [PubMed] [Google Scholar]
  5. Berse B., Dmochowska A., Skrzypek M., Wegleński P., Bates M. A., Weiss R. L. Cloning and characterization of the ornithine carbamoyltransferase gene from Aspergillus nidulans. Gene. 1983 Nov;25(1):109–117. doi: 10.1016/0378-1119(83)90173-7. [DOI] [PubMed] [Google Scholar]
  6. Brown D. W., Yu J. H., Kelkar H. S., Fernandes M., Nesbitt T. C., Keller N. P., Adams T. H., Leonard T. J. Twenty-five coregulated transcripts define a sterigmatocystin gene cluster in Aspergillus nidulans. Proc Natl Acad Sci U S A. 1996 Feb 20;93(4):1418–1422. doi: 10.1073/pnas.93.4.1418. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chater K. F., Bruton C. J., Plaskitt K. A., Buttner M. J., Méndez C., Helmann J. D. The developmental fate of S. coelicolor hyphae depends upon a gene product homologous with the motility sigma factor of B. subtilis. Cell. 1989 Oct 6;59(1):133–143. doi: 10.1016/0092-8674(89)90876-3. [DOI] [PubMed] [Google Scholar]
  8. Choi G. H., Chen B., Nuss D. L. Virus-mediated or transgenic suppression of a G-protein alpha subunit and attenuation of fungal virulence. Proc Natl Acad Sci U S A. 1995 Jan 3;92(1):305–309. doi: 10.1073/pnas.92.1.305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. De Vries L., Mousli M., Wurmser A., Farquhar M. G. GAIP, a protein that specifically interacts with the trimeric G protein G alpha i3, is a member of a protein family with a highly conserved core domain. Proc Natl Acad Sci U S A. 1995 Dec 5;92(25):11916–11920. doi: 10.1073/pnas.92.25.11916. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dohlman H. G., Apaniesk D., Chen Y., Song J., Nusskern D. Inhibition of G-protein signaling by dominant gain-of-function mutations in Sst2p, a pheromone desensitization factor in Saccharomyces cerevisiae. Mol Cell Biol. 1995 Jul;15(7):3635–3643. doi: 10.1128/mcb.15.7.3635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Druey K. M., Blumer K. J., Kang V. H., Kehrl J. H. Inhibition of G-protein-mediated MAP kinase activation by a new mammalian gene family. Nature. 1996 Feb 22;379(6567):742–746. doi: 10.1038/379742a0. [DOI] [PubMed] [Google Scholar]
  12. Hopwood D. A. The Leeuwenhoek lecture, 1987. Towards an understanding of gene switching in Streptomyces, the basis of sporulation and antibiotic production. Proc R Soc Lond B Biol Sci. 1988 Nov 22;235(1279):121–138. doi: 10.1098/rspb.1988.0067. [DOI] [PubMed] [Google Scholar]
  13. Kale S. P., Cary J. W., Bhatnagar D., Bennett J. W. Characterization of experimentally induced, nonaflatoxigenic variant strains of Aspergillus parasiticus. Appl Environ Microbiol. 1996 Sep;62(9):3399–3404. doi: 10.1128/aem.62.9.3399-3404.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. 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]
  15. Koelle M. R., Horvitz H. R. EGL-10 regulates G protein signaling in the C. elegans nervous system and shares a conserved domain with many mammalian proteins. Cell. 1996 Jan 12;84(1):115–125. doi: 10.1016/s0092-8674(00)80998-8. [DOI] [PubMed] [Google Scholar]
  16. Käfer E. Meiotic and mitotic recombination in Aspergillus and its chromosomal aberrations. Adv Genet. 1977;19:33–131. doi: 10.1016/s0065-2660(08)60245-x. [DOI] [PubMed] [Google Scholar]
  17. Lee B. N., Adams T. H. FluG and flbA function interdependently to initiate conidiophore development in Aspergillus nidulans through brlA beta activation. EMBO J. 1996 Jan 15;15(2):299–309. [PMC free article] [PubMed] [Google Scholar]
  18. Lee B. N., Adams T. H. Overexpression of flbA, an early regulator of Aspergillus asexual sporulation, leads to activation of brlA and premature initiation of development. Mol Microbiol. 1994 Oct;14(2):323–334. doi: 10.1111/j.1365-2958.1994.tb01293.x. [DOI] [PubMed] [Google Scholar]
  19. Lee B. N., Adams T. H. The Aspergillus nidulans fluG gene is required for production of an extracellular developmental signal and is related to prokaryotic glutamine synthetase I. Genes Dev. 1994 Mar 15;8(6):641–651. doi: 10.1101/gad.8.6.641. [DOI] [PubMed] [Google Scholar]
  20. PONTECORVO G., ROPER J. A., HEMMONS L. M., MACDONALD K. D., BUFTON A. W. J. The genetics of Aspergillus nidulans. Adv Genet. 1953;5:141–238. doi: 10.1016/s0065-2660(08)60408-3. [DOI] [PubMed] [Google Scholar]
  21. Rens-Domiano S., Hamm H. E. Structural and functional relationships of heterotrimeric G-proteins. FASEB J. 1995 Aug;9(11):1059–1066. doi: 10.1096/fasebj.9.11.7649405. [DOI] [PubMed] [Google Scholar]
  22. Stack M., Rodricks J. V. Method for analysis and chemical confirmation of sterigmatocystin. J Assoc Off Anal Chem. 1971 Jan;54(1):86–90. [PubMed] [Google Scholar]
  23. Trail F., Chang P. K., Cary J., Linz J. E. Structural and functional analysis of the nor-1 gene involved in the biosynthesis of aflatoxins by Aspergillus parasiticus. Appl Environ Microbiol. 1994 Nov;60(11):4078–4085. doi: 10.1128/aem.60.11.4078-4085.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Turner G. E., Borkovich K. A. Identification of a G protein alpha subunit from Neurospora crassa that is a member of the Gi family. J Biol Chem. 1993 Jul 15;268(20):14805–14811. [PubMed] [Google Scholar]
  25. Watson N., Linder M. E., Druey K. M., Kehrl J. H., Blumer K. J. RGS family members: GTPase-activating proteins for heterotrimeric G-protein alpha-subunits. Nature. 1996 Sep 12;383(6596):172–175. doi: 10.1038/383172a0. [DOI] [PubMed] [Google Scholar]
  26. Wieser J., Adams T. H. flbD encodes a Myb-like DNA-binding protein that coordinates initiation of Aspergillus nidulans conidiophore development. Genes Dev. 1995 Feb 15;9(4):491–502. doi: 10.1101/gad.9.4.491. [DOI] [PubMed] [Google Scholar]
  27. Wieser J., Lee B. N., Fondon J. w., 3rd, Adams T. H. Genetic requirements for initiating asexual development in Aspergillus nidulans. Curr Genet. 1994 Dec;27(1):62–69. doi: 10.1007/BF00326580. [DOI] [PubMed] [Google Scholar]
  28. Woloshuk C. P., Foutz K. R., Brewer J. F., Bhatnagar D., Cleveland T. E., Payne G. A. Molecular characterization of aflR, a regulatory locus for aflatoxin biosynthesis. Appl Environ Microbiol. 1994 Jul;60(7):2408–2414. doi: 10.1128/aem.60.7.2408-2414.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Yu J. H., Butchko R. A., Fernandes M., Keller N. P., Leonard T. J., Adams T. H. Conservation of structure and function of the aflatoxin regulatory gene aflR from Aspergillus nidulans and A. flavus. Curr Genet. 1996 May;29(6):549–555. doi: 10.1007/BF02426959. [DOI] [PubMed] [Google Scholar]
  30. Yu J. H., Leonard T. J. Sterigmatocystin biosynthesis in Aspergillus nidulans requires a novel type I polyketide synthase. J Bacteriol. 1995 Aug;177(16):4792–4800. doi: 10.1128/jb.177.16.4792-4800.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Yu J. H., Wieser J., Adams T. H. The Aspergillus FlbA RGS domain protein antagonizes G protein signaling to block proliferation and allow development. EMBO J. 1996 Oct 1;15(19):5184–5190. [PMC free article] [PubMed] [Google Scholar]

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