Analysis of the Relationship between Alternative Respiration and Sterigmatocystin Formation in Aspergillus nidulans

Abstract

Aspergillus nidulans has one gene for alternative oxidase (EC 1.10.3.11). To investigate the relationship between this mitochondrial terminal oxidase and the formation of the mycotoxin sterigmatocystin, the encoding aodA gene was both deleted and overexpressed. Relative to the wild-type, the cyanide-resistant fraction of respiration in the late stationary stage—when sterigmatocystin production occurs—doubled in the overexpressing mutant carrying three aodA gene copies, but decreased to 10% in the deletant. Essentially identical results were obtained regardless whether the cultures were illuminated or protected from light. In contrast, sterigmatocystin yield in the aodA deletant was about half of that in the control when grown in the dark, while aodA overexpression resulted in up to 70% more sterigmatocystin formed, the yield increasing with alternative oxidase activity. Results were quite different when cultures were illuminated: under those conditions, sterigmatocystin volumetric yields were considerably lower, and statistically unvarying, regardless of the presence, absence, or the copy number of aodA. We conclude that the copy number of aodA, and hence, the balance between alternative- and cytochrome C-mediated respiration, appears to correlate with sterigmatocystin production in A. nidulans, albeit only in the absence of light.

Alternative Oxidase: A Potential Target for Controlling Aflatoxin Contamination and Propagation of Aspergillus flavus

“…AOX inhibitors have reportedly prevented M. perniciosa, M. grisea, and Botrytis cinerea spore germination in vitro (Inoue et al, 2012;Barsottini et al, 2019), suggesting that AOX activity is a common feature needed for spore germination-related pathways in these fungi. A previous study reported that on initiation of aflatoxin production, the external carbon source was greatly consumed, thus resulting in aflatoxin biosynthesis from the breakdown of reserve carbon sources such as lipids and fatty acids (Molnár et al, 2018a). AOX overexpression in Podospora anserine increased fatty acid production and decreased 2-oxoglutarate concentrations, suggesting a redirection in cellular metabolism (Bovier et al, 2014).…”

“…Despite a dearth of knowledge of how AOX affects aflatoxin biosynthesis, AOX activity seems to affect sterigmatocystin, the penultimate intermediate in the biosynthesis of aflatoxin B 1 . Molnár et al (2018a) investigated the association between AOX and sterigmatocystin synthesis in A. nidulans by both deleting and overexpressing the gene encoding AOX. Compared with the wild-type, the overexpressing mutant produced up to 70% more sterigmatocystin and the deletion mutant produced 50% less sterigmatocystin when grown in the dark.…”

“…Furthermore, aflatoxin synthesis requires considerable amounts of ATP and NADPH (Yabe and Nakajima, 2004). However, after the initial growth phase, the energy demand of fungal cells is reduced and carbon catabolism is inhibited through oxidative phosphorylation (Cárdenas-Monroy et al, 2017;Molnár et al, 2018a). To maintain cellular homeostasis, fungal cells may rely on alternative respiratory pathways for reoxidation of NADH for aflatoxin synthesis without concomitant ATP production.…”

“…AOX activity is associated with reactive oxygen species (ROS) control, metabolic homeostasis, cellular energy demand, the redox state, and the stress response (Li et al, 2011;Vanlerberghe, 2013). Furthermore, AOX affects mycotoxin production, such as sterigmatocystin, which is the penultimate intermediate in aflatoxin B 1 biosynthesis (Leiter et al, 2016;Molnár et al, 2018a). As AOX is absent in mammals, it has been investigated as a potential drug target for pathogenic fungi (Tudella et al, 2004;Ebiloma et al, 2019).…”

Biochemical characterization and inhibition of the alternative oxidase enzyme from the fungal phytopathogen Moniliophthora perniciosa

“…Interestingly, genetic ablation of AOX activity has led to reduced virulence or survival rates in Cryptococcus neoformans and A. fumigatus 20,21 . Finally, Aspergillus nidulans and A. fumigatus exhibit a correlation between higher AOX activity and higher production of secondary metabolites, such as sterigmatocystin and cephalosporin C 22,23 .…”