A complex metabolic network and its biomarkers regulate laccase production in white-rot fungus Cerrena unicolor 87613

Abstract

Background: White-rot fungi are known to naturally produce high quantities of laccase, which exhibit commendable stability and catalytic efficiency. However, their laccase production does not meet the demands for industrial-scale applications. To address this limitation, it is crucial to optimize the conditions for laccase production. However, the regulatory mechanisms underlying different conditions remain unclear. This knowledge gap hinders the cost-effective application of laccases.

Results: In this study, we utilized transcriptomic and metabolomic data to investigate a promising laccase producer, Cerrena unicolor 87613, cultivated with fructose as the carbon source. Our comprehensive analysis of differentially expressed genes (DEGs) and differentially abundant metabolites (DAMs) aimed to identify changes in cellular processes that could affect laccase production. As a result, we discovered a complex metabolic network primarily involving carbon metabolism and amino acid metabolism, which exhibited contrasting changes between transcription and metabolic patterns. Within this network, we identified five biomarkers, including succinate, serine, methionine, glutamate and reduced glutathione, that played crucial roles in co-determining laccase production levels.

Conclusions: Our study proposed a complex metabolic network and identified key biomarkers that determine the production level of laccase in the commercially promising Cerrena unicolor 87613. These findings not only shed light on the regulatory mechanisms of carbon sources in laccase production, but also provide a theoretical foundation for enhancing laccase production through strategic reprogramming of metabolic pathways, especially related to the citrate cycle and specific amino acid metabolism.

Keywords: Cerrena unicolor; Fructose; Laccase production; Metabolic networks; Regulation mechanism; White rot fungi.

Fig. 1

Laccase production of C. unicolor 87613. To assess the effect of different carbon sources, strains were incubated in PDA media supplied with 20 g/L of glucose, fructose, sucrose, lactose, starch, or dextrin as extra carbon source, respectively. Then, the laccase activity was quantified using ABTS as the substrate

Fig. 2

The transcription pattern of FCd-6 compared to that of FCd-10. A Venn diagram showing the numbers and relationship of all detected transcripts according to setting threshold limit as log2 (fold-change) > 1 or log2(fold-change) < − 1, and P-value < 0.05. B The volcano map of transcription profile of all RNAs in FCd-6 compared to those in FCd-10. The red dots indicate significantly up-regulated genes, whereas the green dots indicate down-regulated genes. The grey dots indicate that genes were not significantly changed. C Gene Ontology (GO) analysis of up-regulated genes (red box) and down-regulated genes (green box) in categories of Biological Process (BP), Cellular Component (CC), and Molecular Function (MF). D KEGG analysis of up-regulated genes (red box) and down-regulated genes (green box) in ten enriched terms

Fig. 3

The metabolic pattern of FCd-6 compared to that of FCd-10. A The volcano map of metabolites profile in FCd-6 compared to those in FCd-10. The increased metabolites were marked in orange dots, whereas the decreased metabolites were marked in blue dots. B Categories of metabolites exhibiting differential abundances. C Number of metabolites with differential abundance. D Metabolic pathways enriched by differentially abundant metabolites (DAMs) using MetaboAnalyst 6.0 online server. The red and green number indicate the increased or decreased DAMs in each pathway, respectively

Fig. 4

Combination analysis of transcriptome and metabolome resulting A eight co-enriched pathways. B six of them constituted a complicated metabolic network with different changes of transcription and metabolic profiles. The left and right box indicated FCd-6 and FCd-10 samples, respectively. The red box indicated significantly up-regulated pattern of transcription, whereas the green box indicated down-regulated pattern of transcription. The yellow box indicated significantly increased pattern of metabolome, whereas the blue box indicated significantly decreased pattern of metabolome

Fig. 5

The alteration of A citrate cycle (TCA) cycle in FCd-6 compared to those FCd-10 as cultivated with fructose. B the ROS levels at FCd-6 and FCd-10 periods. C the biomass of fungal hyphae growing from FCd-2 to FCd-12. D the intensity of succinate measured by LC-MS.MS at FCd-6 and FCd-10 periods. E the dose-dependent induction of laccase production by additive succinate during C. unicolor 87613 cultivation

Fig. 6

The alteration of A cysteine and methionine metabolism pathway in Cd-6 compared to those FCd-10 as cultivated with fructose. B the intensity of serine (Ser) measured by LC-MS.MS at FCd-6 and FCd-10 periods. C the dose-dependent repression of laccase production by additive serine during C. unicolor 87613 cultivation. D the intensity of methionine (Met) measured by LC-MS.MS at FCd-6 and FCd-10 periods. E the dose-dependent repression of laccase production by additive methionine during C. unicolor 87613 cultivation

Fig. 7

The alteration of A glutathione metabolism pathway in Cd-6 compared to those FCd-10 as cultivated with fructose. B the intensity of glutamate (Glu) measured by LC-MS.MS at FCd-6 and FCd-10 periods. C the dose-dependent repression of laccase production by additive glutamate during C. unicolor 87613 cultivation. D the intensity of reduced glutathione (GSH) measured by LC-MS.MS at FCd-6 and FCd-10 periods. E the dose-dependent repression of laccase production by additive GSH during C. unicolor 87613 cultivation

Fig. 8

The illustration of altered laccase production and corresponding intracellular abundance of five metabolic biomarkers (succinate, GSH, glutamate, serine, and methionine) in A FCd-6 versus FCd-10, B GCd-6 versus GCd-10, C FCd-6 versus GCd-6, and D FCd-10 versus GCd-10, respectively. The yellow asterisk indicates a promoting effect on laccase activity, the blue asterisk suggests an inhibitory effect on laccase activity, and the gray asterisk indicates on effect on laccase activity