Carbon catabolite repression (CCR) is regulated by the C2H2-type transcription factor CreA/Cre1 in filamentous fungi. In addition to CreA, three additional factors (CreB, CreC and CreD) have been identified as CCR regulatory factors. CreB and CreC form a deubiquitinating enzyme complex, and disruption of creB promotes amylolytic and biomass-degrading enzymes production in Aspergillus oryzae (1, 2). CreD is an arrestin-like protein that serves as an adaptor of ubiquitin ligase and its target including maltose permease (3). The function of these factors has led us to the hypothesis that proteolytic degradation of the CreA protein is involved in CCR regulation. In this study, we investigated the stability and subcellular localization of FLAG- and GFP-tagged CreA in A. oryzae after the addition of various sugars. The abundance of CreA was dramatically reduced after incubation in maltose and xylose, which stimulated the export of CreA from the nucleus to the cytoplasm. Mutation of a putative nuclear export signal resulted in nuclear retention and significant stabilization of CreA. These results suggest that CreA is rapidly degraded in the cytoplasm after export from the nucleus. The CreA protein level was reduced by disruption of creB and creC, although CreA stability was not affected by disruption of creD. Deletion of the last 40 C-terminal amino acids resulted in remarkable stabilization and increased abundance of CreA, whereas deletion of the last 20 C-terminal amino acids had no apparent effect on CreA stability. This result suggests that the 20 amino acid region located between positions 390 and 409 of CreA is critical for the rapid degradation of CreA. (1) Ichinose and Tanaka et al., 2014, Appl. Microbiol. Biotechnol., 98: 335-343. (2) Ichinose et al., 2018, J. Biosci. Bioeng., 125: 141-147. (3) Tanaka et al., 2017, Appl. Environ. Microbiol., 83: e00592-17.
Full conference title:
- Fungal Genetics Conference 30th (2019)