Spermidine, a kind of polycation and one important member in the polyamine family, is essential for survival in many kinds of organisms and participates in the regulation of cell growth and metabolism. To explore the mechanism by which spermidine regulates ganoderic acids (GAs) biosynthesis in Ganoderma lucidum , the effects of spermidine on GAs and reactive oxygen species (ROS) contents were examined. Our data suggested that spermidine promoted the production of mitochondrial ROS and positively regulated GAs biosynthesis. Further research revealed that spermidine promoted the translation of mitochondrial complexes I and II and subsequently influenced their activity. With the reduction in eukaryotic translation initiation factor 5A (eIF5A) hypusination by over 50% in spermidine synthase gene ( spds ) knockdown strains, the activity of mitochondrial complexes I and II was reduced by nearly 60% and 80%, respectively, and the protein content was reduced by over 50%, suggesting that the effect of spermidine in mitochondrial complexes I and II was mediated through its influence on eIF5A hypusination. Furthermore, after knocking down eIF5A , the deoxyhypusine synthase gene ( dhs ) and the deoxyhypusine hydroxylase gene ( dohh ), mitochondrial ROS level was reduced by nearly 50%, and GAs content was reduced by over 40%, suggesting that eIF5A hypusination contributed to mitochondrial ROS production and GAs biosynthesis. In summary, spermidine maintains mitochondrial ROS homeostasis by regulating the translation and subsequent activity of complexes I and II via eIF5A hypusination and promotes GAs biosynthesis via mitochondrial ROS signaling. The present findings provide new insight into spermidine-mediated biosynthesis of secondary metabolites. IMPORTANCE: Spermidine is necessary for organism survival and is involved in the regulation of various biological processes. However, the specific mechanisms underlying the various physiological functions of spermidine are poorly understood, especially in microorganisms. In this study, we found that spermidine hypusinates eIF5A to promote the production of mitochondrial ROS and subsequently regulate secondary metabolism in microorganisms. Our study provides a better understanding of the mechanism by which spermidine regulates mitochondrial function and provides new insight into spermidine-mediated biosynthesis of secondary metabolites.

Trehalose‐6‐phosphate synthase (TPS) is a key enzyme that participates in trehalose metabolism, which can synthesize trehalose in a two‐step pathway with trehalose phosphatase, but its role in fungi is rarely studied, especially in large basidiomycetes. In this study, the tps gene of Ganoderma lucidum was cloned and named as gltps. And gltps‐silenced strains were constructed by RNA interference. In this study, it is found that the extracellular polysaccharide content increased 1.6–2‐fold, but there was no significant change on intracellular polysaccharide content in gltps‐silenced strains compared with the wild‐type (WT) strain. Furthermore, the cell wall compositions of the gltps‐silenced strains were also altered, which showed that the chitin and β‐1,3‐glucan contents were significantly decreased. Compared with WT, the concentration of chitin decreased by 20%–50% and that of β‐1, 3‐glucan decreased by 15%–30%. The study found that the cells of gltps‐silenced strains were more sensitive to cell wall stress, which might be due to changes in the compounds and structure of the cell wall. These results showed that gltps had an important effect on carbohydrate metabolism of G. lucidum cells.

The activity of mitochondrial pyruvate carrier (MPC) can be modulated to regulate intracellular metabolism under different culture conditions. In Ganoderma lucidum, the role of MPC in regulating carbon sources remains unknown. By knocking down MPC genes (MPC1 and MPC2), this research found that the loss of MPC increased the growth rate of G. lucidum by approximately 30% in a medium with wood chips as a carbon source. Then, cellulase and laccase activities were tested. Endoglucanase and laccase activity increased by approximately 50% and 35% respectively in MPC knockdown mutants compared with that in the WT strain. Finally, the expression levels of genes related to glycolysis were assayed, and the transcription levels of these enzymes were found to be increased by approximately 250% compared with the WT strain. In conclusion, the regulation of intracellular metabolism by MPC provides a new way to improve the use of nondominant carbon sources such as lignocellulose.