Document Type

Article

Subject Area(s)

Biochemistry

Abstract

Background: Filamentous fungi in the genus Aspergillus produce a variety of natural products, including aflatoxin, the most potent naturally occurring carcinogen known. Aflatoxin biosynthesis, one of the most highly characterized secondary metabolic pathways, offers a model system to study secondary metabolism in eukaryotes. To control or customize biosynthesis of natural products we must understand how secondary metabolism integrates into the overall cellular metabolic network. By applying a metabolomics approach we analyzed volatile compounds synthesized by Aspergillus parasiticus in an attempt to define the association of secondary metabolism with other metabolic and cellular processes.

Results: Volatile compounds were examined using solid phase microextraction - gas chromatography/mass spectrometry. In the wild type strain Aspergillus parasiticus SU-1, the largest group of volatiles included compounds derived from catabolism of branched chain amino acids (leucine, isoleucine, and valine); we also identified alcohols, esters, aldehydes, and lipid-derived volatiles. The number and quantity of the volatiles produced depended on media composition, time of incubation, and light-dark status. A block in aflatoxin biosynthesis or disruption of the global regulator veA affected the volatile profile. In addition to its multiple functions in secondary metabolism and development, VeA negatively regulated catabolism of branched chain amino acids and synthesis of ethanol at the transcriptional level thus playing a role in controlling carbon flow within the cell. Finally, we demonstrated that volatiles generated by a veA disruption mutant are part of the complex regulatory machinery that mediates the effects of VeA on asexual conidiation and sclerotia formation.

Conclusions: 1) Volatile profiling provides a rapid, effective, and powerful approach to identify changes in intracellular metabolic networks in filamentous fungi. 2) VeA coordinates the biosynthesis of secondary metabolites with catabolism of branched chain amino acids, alcohol biosynthesis, and b-oxidation of fatty acids. 3) Intracellular chemical development in A. parasiticus is linked to morphological development. 4) Understanding carbon flow through secondary metabolic pathways and catabolism of branched chain amino acids is essential for controlling and customizing production of natural products.

Additional File 1.tiff (439 kB)
Figure S1 - Growth of A. parasiticus strains in YES liquid medium.

Additional File 2.ppt (126 kB)
Figure S2 - SPME-GC/MS headspace gas analysis of selected volatile compounds produced by aspergilli grown in YES medium in the dark for 72 h.

Additional File 3.pdf (52 kB)
Figure S3 - Production of fungal volatiles through pathways of branched chain amino acid catabolism.

Additional File 4.ppt (108 kB)
Figure S4 - Branched chain amino acid-derived volatiles generated by SU-1 grown for 48 h and 72 h in light.

Additional File 5.pdf (21 kB)
Figure S5 - Branched chain amino acid-derived esters detected in A. parasiticus strains.

Additional File 6.pdf (18 kB)
Figure S6 - Amino acid sequence alignment of the putative A. flavus branched chain amino acid aminotransferases AFLA_113800 and AFLA_044190 with yeast BAT1 and BAT2.

Additional File 7.pdf (11 kB)
Figure S7 - Amino acid sequence alignment of the putative A. flavus alcohol dehydrogenase, AFLA_048690, with the yeast alcohol dehydrogenase, ADH1.

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