Identification of polyketide synthase genes required for aspinolides biosynthesis in Trichoderma arundinaceum

Authors: CARDOZA, ROSA - University Of Leon; McCormick, Susan; IZQUIERDO-BUENO, IMMACULADA - University Of Cadiz; MARTÍNEZ-REYES, NATALIA - University Of Leon; LINDO, LAURA - University Of Leon; Brown, Daren; COLLADO, ISIDRO - University Of Cadiz; Proctor, Robert; GUTIERREZ, SANTIAGO - University Of Leon

Submitted to: Applied Microbiology and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/10/2022
Publication Date: 9/27/2022
Citation: Cardoza, R.E., McCormick, S.P., Izquierdo-Bueno, I., Martínez-Reyes, N., Lindo, L., Brown, D.W., Collado, I.G., Proctor, R.H., Gutierrez, S. 2022. Identification of polyketide synthase genes required for aspinolides biosynthesis in Trichoderma arundinaceum. Applied Microbiology and Biotechnology. 106:7153–7171. https://doi.org/10.1007/s00253-022-12182-9.
DOI: https://doi.org/10.1007/s00253-022-12182-9

Interpretive Summary: The fungus Trichoderma arundinaceum is used as a biological control agent for crop diseases caused by other fungi. Its biological control activity stems from the ability of T. arundinaceum to produce metabolites that inhibit growth of crop-disease-causing fungi. However, T. arundinaceum poses a potential threat to food and feed safety because one of the antifungal metabolites it produces, harzianum A, is a member of the trichothecene family of fungal toxins, and many trichothecenes are toxic to humans and domestic animals. Therefore, researchers in Leon, Spain, in collaboration with ARS researchers in Peoria, Illinois, determined whether the ability to produce a group of antifungal metabolites called aspinolides contributes to the biological control activity of T. arundinaceum. The researchers identified genes in T. arundinaceum that are responsible for aspinolide synthesis and showed that blocking aspinolide production did not decrease the ability of T. arundinaceum to inhibit the growth of other fungi, possibly because blocking aspinolide production increased harzianum A production. These findings demonstrate the complex biochemical nature of the antifungal activity of T. arundinaceum. Such knowledge contributes to efforts to make the fungus a safe biological control agent.

Technical Abstract: The fungus Trichoderma arundinaceum exhibits biological control activity against crop diseases caused by other fungi. Two mechanisms that likely contribute to this activity are upregulation of plant defenses and production of two types of antifungal secondary metabolites: the sesquiterpenoid harzianum A (HA) and the polyketide-derived aspinolides. The goal of the current study was to identify aspinolide biosynthetic genes as part of an effort to understand how these metabolites contribute to the biological control activity of T. arundinaceum. Comparative genomics identified two polyketide synthase genes (asp1 and asp2) that occur in T. arundinaceum and Aspergillus ochraceus, which also produces aspinolides. Gene deletion and biochemical analyses in T. arundinaceum indicated that both genes are required for aspinolide production: asp2 for formation of a 10-member lactone ring and asp1 for formation of a butenoyl subsituent at position 8 of the lactone ring. Gene expression and comparative genomics analyses indicated that asp1 and asp2 are located within a gene cluster that occurs in both T. arundinaceum and A. ochraceus. A survey of genome sequences representing 35 phylogenetically diverse Trichoderma species revealed that intact homologs of the cluster occurred in only two other species, which also produced aspinolides. An asp2 mutant inhibited fungal growth more than the wild type, but an asp1 mutant did not, and the greater inhibition by the asp2 mutant coincided with increased HA production. These findings indicate that asp1 and asp2 are aspinolide biosynthetic genes and that loss of either aspinolide or HA production in T. arundinaceum can be accompanied by increased production of the other metabolite(s).