A Gene from with Similarity to of Filamentous Ascomycetes Contributes to Regulating AreA.

Fungal AreA is a key nitrogen metabolism transcription factor in nitrogen metabolism repression (NMR). Studies have shown that there are different ways to regulate AreA activity in yeast and filamentous ascomycetes, but in , how AreA is regulated is unknown. Here, a gene from with similarity to of filamentous ascomycetes was identified. The NmrA interacted with the C-terminal of AreA according to yeast two-hybrid assay. In order to determine the effect of NmrA on the AreA, 2 silenced strains of , with silencing efficiencies of 76% and 78%, were constructed using an RNA interference method. Silencing resulted in a decreased content of AreA. The content of AreA in i-3 and i-48 decreased by approximately 68% and 60%, respectively, compared with that in the WT in the ammonium condition. Under the nitrate culture condition, silencing resulted in a 40% decrease compared with the WT. Silencing also reduced the stability of the AreA protein. When the mycelia were treated with cycloheximide for 6 h, the AreA protein was almost undetectable in the silenced strains, while there was still approximately 80% of the AreA protein in the WT strains. In addition, under the nitrate culture, the content of AreA protein in the nuclei of the WT strains was significantly increased compared with that under the ammonium condition. However, when was silenced, the content of the AreA protein in the nuclei did not change compared with the WT. Compared with the WT, the expression of the glutamine synthetase gene in i-3 and i-48 strains increased by approximately 94% and 88%, respectively, under the ammonium condition, while the expression level of the nitrate reductase gene in i-3 and i-48 strains increased by approximately 100% and 93%, respectively, under the nitrate condition. Finally, silencing inhibited mycelial growth and increased ganoderic acid biosynthesis. Our findings are the first to reveal that a gene from with similarity to the of filamentous ascomycetes contributes to regulating AreA, which provides new insight into how AreA is regulated in .