YPR2 is a regulator of light modulated carbon and secondary metabolism in Trichoderma reesei

Abstract

Background: Filamentous fungi have evolved to succeed in nature by efficient growth and degradation of substrates, but also due to the production of secondary metabolites including mycotoxins. For Trichoderma reesei, as a biotechnological workhorse for homologous and heterologous protein production, secondary metabolite secretion is of particular importance for industrial application. Recent studies revealed an interconnected regulation of enzyme gene expression and carbon metabolism with secondary metabolism.

Results: Here, we investigated gene regulation by YPR2, one out of two transcription factors located within the SOR cluster of T. reesei, which is involved in biosynthesis of sorbicillinoids. Transcriptome analysis showed that YPR2 exerts its major function in constant darkness upon growth on cellulose. Targets (direct and indirect) of YPR2 overlap with induction specific genes as well as with targets of the carbon catabolite repressor CRE1 and a considerable proportion is regulated by photoreceptors as well. Functional category analysis revealed both effects on carbon metabolism and secondary metabolism. Further, we found indications for an involvement of YPR2 in regulation of siderophores. In agreement with transcriptome data, mass spectrometric analyses revealed a broad alteration in metabolite patterns in ∆ypr2. Additionally, YPR2 positively influenced alamethicin levels along with transcript levels of the alamethicin synthase tex1 and is essential for production of orsellinic acid in darkness.

Conclusions: YPR2 is an important regulator balancing secondary metabolism with carbon metabolism in darkness and depending on the carbon source. The function of YPR2 reaches beyond the SOR cluster in which ypr2 is located and happens downstream of carbon catabolite repression mediated by CRE1.

Keywords: Alamethicin; Carbon metabolism; Hypocrea jecorina; Light response; Secondary metabolism; Trichoderma reesei.

Fig. 1

Gene regulation by YPR2 in T. reesei. a Schematic representation of the SOR cluster. Genomic locations are taken from the JGI Trichoderma reesei database v2.0 (https://genome.jgi.doe.gov/Trire2/Trire2.home.html). b Hierarchical clustering of gene regulation patterns in ∆ypr2 compared to wildtype in constant light (LL) and constant darkness (DD) upon growth on cellulose. c Numbers of genes regulated in ∆ypr2 in constant light or constant darkness on cellulose (≥2fold, p-value threshold 0.01). d Genes directly or indirectly regulated by YPR2 in constant light overlapping with gene regulation by YPR2 in constant darkness. The diagram shows the proportion of consistent regulation (upregulation in ∆ypr2 in light and darkness, downregulation in light and darkness) or contrasting regulation (upregulation in light and downregulation in darkness (“up”) or downregulation in light and upregulation in darkness (“down”))

Fig. 2

Schematic representation of functional category analysis. a Funcat analysis of genes up- regulated in ∆ypr2 in darkness. b Funcat analysis of genes downregulated in ∆ypr2 in darkness. For funcat overview in light see Additional file 3: Figure S1

Fig. 3

Comparison of gene regulation by YPR2 in darkness with targets (direct or indirect) of CRE1. Amount of genes regulated in ∆ypr2 in constant darkness compared to wildtype versus those regulated in ∆cre1 in darkness. In ∆cre1 233 genes are upregulated in constant darkness and 244 genes are downregulated in constant darkness [5]. Of the 447 genes regulated by CRE1 in darkness, 62 are consistently upregulated in both mutant strains (light green area) and 58 are consistently downregulated in both mutants. In total, of the 447 genes regulated by in ∆cre1 in darkness, 120 are consistently regulated in ∆ypr2 suggesting a double lock mechanism for these genes

Fig. 4

Secondary metabolite production in ∆ypr2 upon growth on cellulose. a Results from mass spectrometric analysis revealed 6 clusters of regulation patterns. b Box plots show levels within the clusters as normalized to biomass formation. Mostly, biosynthesis level even decrease below wildtype in the dark. For smaller sets (cluster 5) elevated levels were observed in the mutant compared to wildtype. c Abundance of Alamethicine in samples lacking sor5 (TR_73623) and ypr2 (TR_102497) upon growth on minimal media with cellulose as carbon source, relative to QM6a and normalized to the biomass produced under these conditions. Errorbars indicate standard deviations of at least two biological replicates