Dual functions of AreA, a GATA transcription factor, on influencing ganoderic acid biosynthesis in Ganoderma lucidum

Abstract

Summary Nitrogen metabolism repression (NMR) has been well studied in filamentous fungi, but the molecular mechanism of its effects on fungal secondary metabolism has been generally unexplored. Ganoderic acid (GA) biosynthesis in Ganoderma lucidum differs between ammonia and nitrate nitrogen sources. To explain the functions of NMR in secondary metabolism, AreA, which is a core transcription factor of NMR, was characterized in G. lucidum. The transcription level of AreA was dramatically increased (approximately 4.5‐folds), with the nitrate as the sole nitrogen source, compared with that with ammonia as the source. In addition, the expression of related genes involved in NMR was changed (upregulated of MeaB and downregulated of Nmr and GlnA) when AreA was knockdown. Yeast one‐hybrid and electrophoretic mobility shift assay results showed that AreA could directly bind to the promoter of fps (encoding farnesyl‐diphosphate synthase) to activate its expression. However, GA biosynthesis was increased (27% in the ammonia source and 77% in the nitrate source) in AreAi mutant strains versus that in control strains. These results showed that another important factor must participate in regulating GA biosynthesis other than the direct activation of AreA. Furthermore, we found that the content of nitric oxide (NO) was increased approximately 2.7‐folds in the nitrate source compared with that in the ammonia. By adding the NO donor (SNP) or scavenger (cPTIO) and using NR‐silenced or NR‐overexpressed strains, we found that there was a negative correlation between the NO contents and GA biosynthesis. NO generated by nitrate reductase (NR) during the nitrogen utilization burst and could negatively influence GA biosynthesis. As a global transcription factor, AreA could also regulate the expression of NR. Our studies provide novel insight into the dual functions of AreA in GA biosynthesis during nitrogen assimilation.

Dual sgRNA‐directed gene deletion in basidiomycete Ganoderma lucidum using the CRISPR/Cas9 system

“…Genetic transformation systems, including Agrobacterium tumefacien ‐mediated and polyethylene glycol (PEG)‐mediated transformations, have been developed in G. lucidum (Shi et al , ; Xu et al , ; Xu and Zhong, ) and have been successfully used to enhance the production of bioactive compounds, such as ganoderic acids and polysaccharides (Zhou et al , ; Li et al , ; Zhang et al , , ; Fei et al , ; Xu et al , ). Gene silencing systems have also been used for the partial suppression of genes of interest in G. lucidum (Mu et al , ; Liu et al , ; Zhu et al , ). So far, however, targeted gene deletion has not been established in Ganoderma , although it would be a powerful tool for G. lucidum research in the post‐genomic era.…”

Regulation of glutamine synthetase activity by transcriptional and posttranslational modifications negatively influences ganoderic acid biosynthesis in Ganoderma lucidum

“…These results suggest that the decrease in GS activity was not due to the GS content. Based on the findings in our previous study, we wondered whether NO, which is an active gas that could burst in nitrate cultivation (Zhu et al ., 2019), affects GS activity via posttranslational regulation. We examined the effect of NO on GS activity in fermentation as our previous study has revealed that the addition of NO donor (SNP) or extruder (PTIO) could increase or decrease the intracellular NO content in the mycelia of G .…”

“…We examined the effect of NO on GS activity in fermentation as our previous study has revealed that the addition of NO donor (SNP) or extruder (PTIO) could increase or decrease the intracellular NO content in the mycelia of G . lucidum (Zhu et al ., 2019). SNP addition decreased GS activity by 28.5% under ammonia conditions, whereas PITO increased GS activity by 15.3% under nitrate conditions compared with the control (Fig.…”

“…It seems that the regulation of GS activity is a complex process. As NO bursts when mycelia are cultured with nitrate (Zhu et al ., 2019), we wondered whether NO might play important roles in regulating GS activity.…”

“…Nitrogen sources are important factors that have been reported to significantly influence GA biosynthesis (Hsieh and Yang, 2004; Zhao et al ., 2011). Many studies have shown that AreA, a key transcription factor, can respond to environmental nitrogen source conditions by activating the expression of nitrogen‐related genes (Wilson and Arst Jr., 1998; Berger et al ., 2006; Zhu et al ., 2019). Our previous study also showed that in the cultivation of nonpreferred nitrogen source, AreA was activated, and it could then promote nitrogen utilization and secondary metabolism by binding to the promoters of related genes including nitrate reductase‐coding gene and genes involved in secondary metabolism (Zhu et al ., 2019).…”