Project description:To investigate the roles of epigenetic regulator homologs, heterochromatin protein 1 (HepA) and a putative methyltransferase (LaeA), in the citric acid produciton by Aspergillus luchuensis mut. kawachii, we constructed the ΔhepA and ΔlaeA strains. We then performed gene expression profiling analysis using data obtained from RNA-seq of Aspergillus luchuensis mut. kawachii control, ΔhepA, and ΔlaeA strains.
Project description:We report that the RNA-binding protein NrdA has a significant role on the global mRNA expression in the white koji fungus, Aspergillus luchuensis mut. kawachii. We constructed the nrdA conditional expression strain using the Tet-On system in A. kawachii. Downregulation of nrdA caused severe growth defect, indicating that NrdA is essential for proliferation of A. kawachii. Parallel RNA-seq and RNA immunoprecipitation (RIP)-seq analysis identified potential NrdA-interacted transcripts, corresponding to 32% of predicted protein coding genes of A. kawachii. Subsequent gene ontology analysis suggested that overexpression of NrdA affects a variety of metabolic activity including secondary metabolite production.
Project description:LaeA, a putative methyltransferase, is a global regulator for metabolic and development process in filamentous fungi. We characterized the laeA homologous genes in the white koji fungus, Aspergillus luchuensis mut. kawachii (A. kawachii) to examine their role in citric acid production. The ΔlaeA strain showed a significant reduction in the citric acid productivity. The cap-analysis gene expression (CAGE) revealed the laeA is required for the gene expression of a putative citrate exporter encoding cexA, which has a critical role for the citric acid production. The deficient citric acid productivity of the ΔlaeA strain was remedied by overexpression of cexA to a comparative level to that of the cexA overexpressing ΔcexA strain. In addition, chromatin immunoprecipitation coupled with quantitative PCR (ChIP-qPCR) analysis indicated that LaeA regulates gene expression of the citrate exporter encoding cexA gene via histone H3K4 and histone H3K9 methylation levels. These results indicate that LaeA is involved in the citric acid production through epigenetic regulation of cexA in A. kawachii.
Project description:Aspergillus luchuensis is a kuro (black) koji mold that have been used as starch degrader for awamori and shochu making industries in Japan. In this study, we investigated the effect of ion beam irradiation on the A. luchuensis RIB2601 and successfully obtained a high starch degrading mutant strain U1. Strain U1 showed reduced growth rate, whereas showed higher α-amylase, glucoamylase, and α-glucosidase activities per mycelial growth than those of wild type strain both in the agar plate and rice koji. In addition, strain U1 showed higher N-acetylglucosamine content in the cell wall and showed higher sensitivity to calcofluor white, indicating deficient cell wall integrity. Interestingly, the analysis of secreted protein showed higher expression of acid labile α-amylase (AmyA) and glucoamylase (GlaA) in strain U1 although the real-time-RT-PCR analysis indicated that the transcriptional levels of the amyA and glaA genes are not changed significantly. These results suggested that the high amylolytic activity of strain U1 is attributed to the high AmyA and GlaA expression level, but it is not through transcriptional regulation of the amyA and glaA genes. Furthermore, the RNA-seq analysis indicated that strain U1 shows transcriptional change of at least 604 genes related to oxidation-reduction, transport, and glucosamine-containing compound metabolic processes, which may be involved in the deficient cell wall integrity of strain U1.
