Characterization of the promoter of the nitrate transporter-encoding gene nrtA in Aspergillus nidulans

Aspergillus nidulans nrtA encodes a nitrate transporter that plays an important role in the NO3- assimilatory process. Many studies have focused on protein functions rather than gene regulation. The knowledge of nrtANO3- uptake process, particularly in the regulation mechanism of transcription factors AreA and NirA on nrtA transcription, is very limited. Herein, we investigated the transcriptional regulation of nrtA in response to various N-sources in detail and characterized the promoter activity of nrtA. We confirmed that nrtA was induced by NO3- and repressed by preferred N-sources. Additionally, for the first time, we found that the transcription of nrtA increased under N-starvation conditions. AreA mediates nrtA transcription under both NO3- and N-starvation conditions, while NirA is effective only under NO3- conditions. All of the proposed AreA and NirA binding sites in the promoter region were capable of binding to their corresponding transcription factors in vitro. In vivo, all of the NirA binding sites showed regulation activities, but to AreA, only several of the initiation-codon-proximal binding sites participated in nrtA transcription. Moreover, the active binding sites contributed in different degrees of regulation strength to nrtA transcription, which is unrelated to the distance between the binding sites and initiation codon. These results provided an extensive map of nrtA promoter, defining the functional regulatory elements of A. nidulans nrtA.