Expression profiles of amylolytic genes in AmyR and CreA transcription factor deletion mutants of the black koji mold Aspergillus luchuensis

The black koji mold, Aspergillus luchuensis, which belongs to Aspergillus section Nigri, is used for the production of traditional Japanese spirits (shochu) mainly in the southern districts of Japan. This mold is known to produce amylolytic enzymes essential for shochu production; however, mechanisms regulating amylolytic gene expression in A. luchuensis have not been studied in as much detail as those in the yellow koji mold, Aspergillus oryzae. Here, we examined the gene expression profiles of deletion mutants of transcription factors orthologous to A. oryzae AmyR and CreA in A. luchuensis. A. luchuensis produces acid-unstable (AmyA) and acid-stable (AsaA) α-amylases. AmyA production and amyA gene expression were not influenced by amyR or creA deletion, indicating that amyA was constitutively expressed. In contrast, asaA gene expression was significantly down- and upregulated upon deletion of amyR and creA, respectively. Furthermore, the glaA and agdA genes (encoding glucoamylase and α-glucosidase, respectively) showed expression profiles similar to those of asaA. Thus, genes that play pivotal roles in starch saccharification, asaA, glaA, and agdA, were found to be regulated by AmyR and CreA. Moreover, despite previous reports on AsaA being only produced in solid-state culture, deletion of the ortholog of A. oryzae flbC, which is involved in the expression of the solid-state culture-specific genes, did not affect AsaA α-amylase activity, suggesting that FlbC was not associated with asaA expression.