Rui-Qi Li

Asperpyridone A represents an unusual class of pyridone alkaloids with demonstrated potential for hypoglycemic activity, primarily by promoting glucose consumption in HepG2 cells. Trichodin A, initially isolated from the marine fungus Trichoderma sp. strain MF106, exhibits notable antibiotic activities against Staphylococcus epidermidis. Despite their pharmacological significance, the regulatory mechanisms governing their biosynthesis have remained elusive. In this investigation, we initiated the activation of a latent gene cluster, denoted as “top”, through the overexpression of the Zn₂Cys₆ transcription factor TopC in Tolypocladium ophioglossoides. The activation of the top cluster led to the biosynthesis of asperpyridone A, pyridoxatin, and trichodin A. Our study also elucidated that the regulator TopC exerts precise control over the biosynthesis of asperpyridone A and trichodin A through the detection of protein–nucleic acid interactions. Moreover, by complementing these findings with gene deletions involving topA and topH, we proposed a comprehensive biosynthesis pathway for asperpyridone A and trichodin A in T. ophioglossoides.

As a potential protein kinase C inhibitor, the fungus metabolite balanol has become more attractive in recent decades. In our previous work, we revealed its biosynthetic pathway through overexpression of the cluster-situated regulator gene blnR in Chinese herb fungus Tolypocladium ophioglossoides. However, information on the regulation of blnR is still largely unknown. In this study, we further investigated the regulation of balanol biosynthesis by BlnR through the analysis of affinity binding using EMSA and RNA-seq analysis. The results showed that BlnR positively regulates balanol biosynthesis through binding to all promoters of bln gene members, including its own promoter. Microscopic observation revealed blnR overexpression also affected spore development and hypha growth. Furthermore, RNA-seq analysis suggested that BlnR can regulate other genes outside of the balanol biosynthetic gene cluster, including those involved in conidiospore development. Finally, balanol production was further improved to 2187.39 mg/L using the optimized medium through statistical optimization based on response surface methodology.