The pleiotropic phenotype of FlbA of Aspergillus niger is explained in part by the activity of seven of its downstream-regulated transcription factors.
Aspergillus niger fulfills an important role in nature in nutrient cycling and is a main cell factory for the industrial production of metabolites and enzymes. Inactivation of flbA in A. niger results in thinner cell walls, increased cell lysis, abolished sporulation, and an increased secretome complexity. A total of 36 transcription factor genes are differentially expressed in ?flbA. Here, seven of these genes (abaA, aslA, aslB, azf1, htfA, nosA, and srbA) were inactivated. Inactivation of each of the genes affected sporulation. Moreover, inactivation of each of the genes impacted protein secretion with the exception of abaA. The impact on secretion was strongest in the case of ?aslA and ?aslB that showed increased pepsin, cellulase, and amylase activity. Biomass formed on agar plates was reduced when either abaA, aslA, nosA, or srbA were inactivated. In contrast, biomass of ?aslA and ?aslB was higher in liquid shaken cultures. The ?aslA and ?htfA strains showed reduced resistance to H2O2, while ?abaA showed increased resistance to cell wall perturbing agents. By contrast, AslA, AslB, HtfA, Azf1, NosA, and SrbA were shown to be important for cell wall integrity. Together, the seven genes downstream of FlbA impact biomass formation, sporulation, protein secretion, and stress resistance in A. niger, and thereby explain part of the pleiotropic phenotype of the ?flbA strain. Overall design: Total RNA of A. niger was isolated from 16 h TM-G and 4 h MM-X liquid shaking cultures. Mycelium of biological triplicates of liquid shaken cultures of A. niger strains MA234.1, ?aslA, and ?aslB were homogenized in a Tissue Lyzer II (Qiagen) under liquid nitrogen. Total RNA was isolated using TRIzol reagent (Invitrogen, www.thermofisher.com). The RNA was purified using the Rnase Plant Mini Kit (Qiagen) and quantified with a Nanodrop ND-1000 (Thermo Scientific, www.thermofisher.com). Sequencing of RNA was performed with Illumina NextSeq2000 2x50bp paired-end technology (Utrecht Sequencing Facility; useq.nl).