Identification and Characterization of the Asperthecin Gene Cluster of Aspergillus nidulans

Abstract

The sequencing of Aspergillus genomes has revealed that the products of a large number of secondary metabolism pathways have not yet been identified. This is probably because many secondary metabolite gene clusters are not expressed under normal laboratory culture conditions. It is, therefore, important to discover conditions or regulatory factors that can induce the expression of these genes. We report that the deletion of sumO, the gene that encodes the small ubiquitin-like protein SUMO in A. nidulans, caused a dramatic increase in the production of the secondary metabolite asperthecin and a decrease in the synthesis of austinol/ dehydroaustinol and sterigmatocystin. The overproduction of asperthecin in the sumO deletion mutant has allowed us, through a series of targeted deletions, to identify the genes required for asperthecin synthesis. The asperthecin biosynthesis genes are clustered and include genes encoding an iterative type I polyketide synthase, a hydrolase, and a monooxygenase. The identification of these genes allows us to propose a biosynthetic pathway for asperthecin.Secondary metabolites are a remarkably rich source of medically useful compounds. A survey of the literature on 1,500 secondary metabolites isolated and characterized between 1993 and 2001 revealed that over half of these compounds have antibacterial, antifungal, or antitumor activity (19). Fungal secondary metabolites, in particular, include a number of important compounds, such as penicillin, cephalosporin, the antihypercholesterolemic agent lovastatin and other statins, and immunosuppressants such as cyclosporine, as well as antifungals (reviewed in references 3, 13, 16, and 19). Other fungal secondary metabolites are important not for their benefits but rather for the problems they cause. For example, the carcinogenic toxins aflatoxin and sterigmatocystin are produced by members of the genus Aspergillus (see references 7, 28, and 29 and earlier references therein; reviewed additionally in references 1 and 26).The sequencing of fungal genomes has revealed several important things about fungal secondary metabolism. First, as was suspected from the results of previous work (13), the genes of pathways that produce particular secondary metabolites are often clustered together (10, 14, 18). Second, fungi have many more secondary metabolism pathways than was previously thought. Analyses of the A. nidulans genome, for example, indicate that A. nidulans has 50 clusters that are predicted to synthesize secondary metabolites (27 polyketides, 14 nonribosomal peptides, 6 fatty acids, 1 terpene, and 2 indole alkaloids) (3, 18). Our own genomic analyses suggest that this number may be a slight overestimate (because more than one polyketide synthase [PKS] and/or nonribosomal peptide synthetase may be involved in a single pathway), but clearly A. nidulans has the ability to synthesize many secondary metabolites. Only a limited number of secondary metabolites in A. nidulans have been identified (aspyridones A and B [from a single pathway], aspoqui...

Elucidation of cladofulvin biosynthesis reveals a cytochrome P450 monooxygenase required for anthraquinone dimerization

“…The nrPKSs MdpG, ACAS, EncA, AptA, and AdaA involved in the first step of monodictyphenone, atrochrysone 2, endocrocin 3, asperthecin, and TAN-1612 biosynthesis, respectively, do not contain any canonical thiolesterase (TE) or Claisen cyclase domains (7,14,(20)(21)(22). Instead, the metallo-β-lactamase-type TEs MdpF, ACTE, EncB, AptB, and AdaB, respectively, are required for closure and release of each nascent polyketide.…”

Dehydrogenase GRD1 Represents a Novel Component of the Cellulase Regulon in Trichoderma reesei (Hypocrea jecorina)

“…The variable orientations yet comparable compositions of genes found in the genomic area around grd1 in these fungi, including a polyketide synthase, suggest the presence of a cluster involved in secondary metabolism. The aspergilli are known for their potent and well-studied secondary metabolism (70,72), and in many cases, genes involved in this process are clustered (2,8,9,63). In contrast, N. crassa was not even known to possess secondary metabolism, except for carotene and melanin biosynthesis, prior to sequencing of the genome (18).…”

Classification, Prediction, and Verification of the Regioselectivity of Fungal Polyketide Synthase Product Template Domains

“…Together, the C2-C7, C4-C19, and the C6-C11 PT domains represent the three known fungal "cyclases" reconstituted so far. It is interesting to note that the AptA PT domain functions efficiently on a nonaketide backbone, while asperthecin was predicted to derive from an octaketide backbone (25). Thus the PT domains may also be functional when paired with minimal PKSs that are of different chain length specificities.…”

“…The commonly observed C6-C11 F-mode cyclization is exemplified by the model product emodin and related compounds (25)(26)(27)(28) (Fig. 1B).…”

“…1B). NRPKSs that synthesize C6-C11 cyclized aromatic polyketides include AptA and AN0150 involved in the synthesis of asperthecin and emodin in Aspergillus nidulans, respectively (25,26), as well as ACAS that synthesizes atrochrysone in Aspergillus terreus (Fig. 1A) (29).…”