title: Enhanced production of bovine chymosin by autophagy deficiency in the filamentous fungus Aspergillus oryzae.
abstract: Aspergillus oryzae has been utilized as a host for heterologous protein production because of its high protein secretory capacity and food-safety properties. However, A. oryzae often produces lower-than-expected yields of target heterologous proteins due to various underlying mechanisms, including degradation processes such as autophagy, which may be a significant bottleneck for protein production. In the present study, we examined the production of heterologous protein in several autophagy (Aoatg) gene disruptants of A. oryzae. We transformed A. oryzae gene disruptants of Aoatg1, Aoatg13, Aoatg4, Aoatg8, or Aoatg15, with a bovine chymosin (CHY) expression construct and found that the production levels of CHY increased up to three fold compared to the control strain. Notably, however, conidia formation by the Aoatg gene disruptants was significantly reduced. As large amounts of conidia are necessary for inoculating large-scale cultures, we also constructed Aoatg gene-conditional expression strains in which the promoter region of the Aoatg gene was replaced with the thiamine-controllable thiA promoter. Conidiation by the resultant transformants was clearly enhanced in the absence of thiamine, while autophagy remained repressed in the presence of thiamine. Moreover, these transformants displayed increased CHY productivity, which was comparable to that of the Aoatg gene disruptants. Consequently, we succeeded in the construction of A. oryzae strains capable of producing high levels of CHY due to defects in autophagy. Our finding suggests that the conditional regulation of autophagy is an effective method for increasing heterologous protein production in A. oryzae.
authors: Department of Biotechnology, The University of Tokyo, Tokyo, Japan.
authors: Jaewoo
authors: J
authors: Yoon
authors: Takashi
authors: T
authors: Kikuma
authors: Jun-ichi
authors: J
authors: Maruyama
authors: Katsuhiko
authors: K
authors: Kitamoto
mesh_terms: D000818:Animals; D001236:Aspergillus oryzae / Q000235:genetics* / Q000378:metabolism; D001343:Autophagy / Q000235:genetics*; D002417:Cattle; D012085:Chymosin / Q000096:biosynthesis* / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D005818:Genetic Engineering; D005822:Genetic Vectors; D008869:Microtubule-Associated Proteins / Q000172:deficiency / Q000235:genetics*; D011401:Promoter Regions, Genetic; D013172:Spores, Fungal / Q000235:genetics* / Q000378:metabolism; D019076:Transgenes
chemical_list: D005656:Fungal Proteins; D008869:Microtubule-Associated Proteins; D012085:Chymosin
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