Characterization and functional analysis of ERAD-related AAA+ ATPase Cdc48 in Aspergillus oryzae |
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| Affiliation: | 1. The United Graduate School of Agricultural Sciences, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan;2. Kirishima Shuzo Co., Ltd., 4-28-1 Shimokawahigashi, Miyakonojo, Miyazaki 885-8588, Japan;3. Faculty of Agriculture, University of the Ryukyus, 1 Senbaru, Nishihara-cho, Okinawa 903-0213, Japan;4. National Research Institute of Brewing, 3-7-1 Kagamiyama, Higashi-Hiroshima, Hiroshima 739-0046, Japan;5. Faculty of Agriculture, Kagoshima University, 1-21-24 Korimoto, Kagoshima 890-0065, Japan;1. Department of Bioscience and Biotechnology, Faculty of Agriculture, Kyushu University, 744 Motooka, Nishi-ku, Fukuoka 819-0395, Japan;2. ASPEX Division, Research Centre, Asahi Glass Co., Ltd., 1150 Hazawa-cho, Yokohama, Kanagawa 221-8755, Japan;3. Department of Applied Biological Science, Faculty of Agriculture, Kagawa University, 2393 Ikenobe, Miki-cho, Kita-gun, Kagawa 761-0795, Japan;1. Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China;2. Fungal Physiology Research Group, CBS-KNAW Fungal Biodiversity Centre, Uppsalalaan 8, 3584 CT, Utrecht, The Netherlands;3. Key Laboratory of Insect Developmental and Evolutionary Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China;4. State Key Laboratory of Microbial Technology, Shandong University, Jinan, Shandong 250100, China;1. Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, 606-8502, Japan;2. Environmental Science Graduate School, The University of Shiga Prefecture, Hikone, Shiga, 522-8533, Japan;1. Microbiology & Fermentation Technology Department, CSIR-Central Food Technological Research Institute, Mysuru, Karnataka, 570020, India;2. AcSIR-Academy of Scientific & Innovative Research, Ghaziabad, UP, 201002, India |
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| Abstract: | Aspergillus oryzae can secrete large amounts of enzymes. However, the production of abundant secretory proteins triggers the unfolded protein response (UPR) in the endoplasmic reticulum (ER), and it is not clear how ER-associated protein degradation (ERAD) contributes to bulk protein production in A. oryzae. Here we identified AoCdc48, the sole A. oryzae ortholog of Saccharomyces cerevisiae AAA+ ATPase Cdc48, a component of the ERAD machinery. We found that AoCdc48 localizes in both nuclei and cytoplasm. Generation of an Aocdc48 conditional mutant showed that Aocdc48 repression leads to reduced cell growth and aberrant hyphal morphology. When Aocdc48-repressed cells were cultured on starch-containing plates, the α-amylase-encoding gene amyB was about 1.3-fold higher expressed. Indeed, a halo produced by secreted amylase was seen on potato starch-containing plates even when there was almost no growth under Aocdc48 repression. Fluorescence microscopy revealed that although AmyB seemed to be secreted, various organelle distributions were aberrant in Aocdc48-repressed cells. We found that D1 AAA domain is crucial for cell viability. Finally, we show that Aocdc48-overexpression also causes defects of cell growth, colonial morphology and conidial formation. Collectively, our results suggest that AoCdc48 is essential for growth and organelle distribution but dispensable for amylase secretion. |
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| Keywords: | AAA ATPase Amylase Cdc48 ERAD |
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