Overexpression of llm1 Affects the Synthesis of Secondary Metabolites of Aspergillus cristatus

Abstract

Putative methyltransferases are thought to be involved in the regulation of secondary metabolites in filamentous fungi. Here, we report the effects of overexpression of a predicted LaeA-like methyltransferase gene llm1 on the synthesis of secondary metabolites in Aspergillus cristatus. Our results revealed that overexpression of the gene llm1 in A. cristatus significantly hindered the production of conidia and enhanced sexual development, and reduced oxidative tolerance to hydrogen peroxide. Compared with the wild-type, the metabolic profile of the overexpression transformant was distinct, and the contents of multiple secondary metabolites were markedly increased, mainly including terpenoids and flavonoids, such as (S)-olEuropeic acid, gibberellin A62, gibberellin A95, ovalitenone, PD 98059, and 1-isomangostin. A total of 600 significantly differentially expressed genes (DEGs) were identified utilizing transcriptome sequencing, and the DEGs were predominantly enriched in transmembrane transport and secondary metabolism-related biological processes. In summary, the strategy of overexpressing global secondary metabolite regulators successfully activated the expression of secondary metabolite gene clusters, and the numerous secondary metabolites were greatly strengthened in A. cristatus. This study provides new insights into the in-depth exploitation and utilization of novel secondary metabolites of A. cristatus.

Metabolomic profiles of the liquid state fermentation in co-culture of Eurotium amstelodami and Bacillus licheniformis

“…Eurotium amstelodami GZ23, B. licheniformis GZ241, B. subtilis GZ237, and B. amyloliquefaciens GZ121 were isolated from Maotai-flavor Daqu produced in Renhuai City, Guizhou Province, China. According to our previous study ( Wang Y. et al, 2022 ), E. amstelodami was grown on high-concentration sodium chloride MYA solid medium (malt extract 20 g, yeast extract powder 5 g, agar powder 15 g, sucrose 30 g, NaCl 170 g and water 1,000 ml) at 37°C to induce a conidia production, the appropriate concentration of conidia of E. amstelodami was adjusted to 1 × 10 6 spore/mL by a blood count plate before inoculation. Bacillus species were inoculated in 150 ml LB liquid medium (Lysogeny Broth: 10 g/l NaCl, 5 g/l yeast extract, 10 g/l tryptone) at 37°C and 180 rpm/min for 12 h, and the cell concentration was adjusted to 1 × 10 8 CFU/ml.…”

“…Analysis was performed following chromatographic conditions: phase A: 95% water +5% acetonitrile (0.1% formic acid in water), phase B: 47.5% acetonitrile +47.5% isopropanol +5% water (0.1% formic acid in water), t : 0–5 min 100% A, 0.5–2.5 min 75% A, 2.5–13 min 0% A, 13–16 min 0%A. We have previously reported the optimization of the UHPLC conditions and mass spectrometry parameters ( Wang Y. et al, 2022 ). Further, the raw data were imported into the metabolomics software, Progenesis QI (Waters Corporation, Milford, United States), for processing and identification of characteristic peaks.…”