Target of Rapamycin Mediated Ornithine Decarboxylase Antizyme Modulate Intracellular Putrescine and Ganoderic Acid Content in Ganoderma lucidum

Abstract

ABSTRACT Putrescine (Put) has been shown to play an important regulatory role in cell growth in organisms. As the primary center regulating the homeostasis of polyamine (PA) content, ornithine decarboxylase antizyme (AZ) can regulate PA content through feedback. Nevertheless, the regulatory mechanism of Put is poorly understood in fungi. Here, our analysis showed that GlAZ had a modulate effect on intracellular Put content by interacting with ornithine decarboxylase (ODC) proteins and reducing its intracellular protein levels. In addition, GlAZ upregulated the metabolic pathway of ganoderic acid (GA) biosynthesis in Ganoderma lucidum by modulating the intracellular Put content. However, a target of rapamycin (TOR) was found to promote the accumulation of intracellular Put after the GlTOR inhibitor Rap was added exogenously, and unbiased analyses demonstrated that GlTOR may promote Put production through its inhibitory effect on the level of GlAZ protein in GlTOR-GlAZ-cosilenced strains. The effect of TOR on fungal secondary metabolism was further explored, and the content of GA in the GlTOR-silenced strain after the exogenous addition of the inhibitor Rap was significantly increased compared with that in the untreated wild-type (WT) strain. Silencing of TOR in the GlTOR-silenced strains caused an increase in GA content, which returned to the WT state after replenishing Put. Moreover, the content of GA in GlTOR-GlAZ-cosilenced strains was also not different from that in the WT strain. Consequently, these results strongly indicate that GlTOR affects G. lucidum GA biosynthesis via GlAZ. IMPORTANCE Research on antizyme (AZ) in fungi has focused on the mechanism by which AZ inhibits ornithine decarboxylase (ODC). Moreover, there are existing reports on the regulation of AZ protein translation by TOR. However, little is known about the mechanisms that influence AZ in fungal secondary metabolism. Here, both intracellular Put content and GA biosynthesis in G. lucidum were shown to be regulated through protein interactions between GlAZ and GlODC. Furthermore, exploration of upstream regulators of GlAZ suggested that GlAZ was regulated by the upstream protein GlTOR, which affected intracellular Put levels and ganoderic acid (GA) biosynthesis. The results of our work contribute to the understanding of the upstream regulation of Put and provide new insights into PA regulatory systems and secondary metabolism in fungi.

Keywords

• Ganoderma lucidum
• ornithine decarboxylase antizyme
• putrescine
• ODC
• TOR
• secondary metabolic