Exploration and Characterization of Hypoxia-inducible Endogenous Promoters in Aspergillus Niger

Aspergillus niger is widely used for the efficient production of organic acids and enzyme preparations. However, this organism lacks basic genetic elements for dynamic control, especially inducible promoters that can respond to specific environmental signals. Since these are desirable for better adaptation of fermentation to large-scale industrial production, herein, we have identified the two first hypoxia-inducible promoters in A. niger, PsrbB and PfhbA. Their performance under high or low oxygen conditions was monitored using two reporter proteins, green fluorescent protein (EGFP) and β-glucuronidase (GUS). For comparison, basal expression of the general strong promoter PgpdA was lower than PsrbB but higher than PfhbA. However, under hypoxia, both promoters showed higher expression than under hyperoxia, and these values were also higher than those observed for PgpdA. For PsrbB, strength under hypoxia was ~2-3 times higher than under hyperoxia (for PfhbA, 3-9 times higher) and ~2.5-5 times higher than for PgpdA (for PfhbA, 2-3 times higher). Promoter truncation analysis showed that the PsrbB fragment -1024 to -588 bp is the core region that determines hypoxia response. KEY POINTS: The first identification of two hypoxia-inducible promoters in A. niger is a promising tool for modulation of target genes under hypoxia. Two reporter genes revealed a different activity and responsiveness to hypoxia of PfhbA and PsrbB promoters, which is relevant for the development of dynamic metabolic regulation of A. niger fermentation. PsrbB promoter truncation and bioinformatics analysis is the foundation for further research.