Analysis of kexin deletion mutant of Aspergillus oryzae.

• Project/Area Number: 16580051
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Applied microbiology
• Research Institution: Tohoku University
• Principal Investigator: YAMAGATA Youhei Tohoku University, Graduate School of Agricultural Sensing, Assistant Professor, 大学院・農学研究科, 助手 (40230338)
• Co-Investigator(Kenkyū-buntansha): NAKAJIMA Tasuku Tohoku University, Graduate School of Agriculture, Science Professor, 大学院・農学研究科, 教授 (20091720)
• Project Period (FY): 2004 – 2005
• Project Status: Completed (Fiscal Year 2005)
• Budget Amount: ¥3,300,000 (Direct Cost: ¥3,300,000); Fiscal Year 2005: ¥1,700,000 (Direct Cost: ¥1,700,000); Fiscal Year 2004: ¥1,600,000 (Direct Cost: ¥1,600,000)
• Keywords: kexin / Aspergillus oryzae / glucan / cell wall / kexB / kexin / プロセッシングプロテアーゼ / α-グルカン / キチン / 細胞壁合成 / cell integrity経路 / mpkA / キチン合成 / グルカン合成

Research Abstract

The deletion mutant of kexB gene encoding a processing protease, kexin in Aspergillus oryzae showed abnormal phenotype in cell morphology. As a result of transcriptome analysis of the mutant fungi, the deletion caused abnormally higher transcription of cell-wall synthesis related enzyme genes, such as chsB and chsC encoding chitin syntheses and gelB encoding a glucan transferase. As the morphological imperfection and abnormal transcription of the genes were suppressed with high osmolarity, it was thought the destruction of the kexB gene caused abnormality of cell integrity pass way. Actually, it was confirmed that A.oryzae mpkA gene that was the ortholog gene of yeast mpk1 was over translated and the resultant protein of MpkA was phosphorylated. The phenomenon was suppressed by high osmolarity stress. These results indicate that the abnormal transcription of cell-wall synthesis related genes and morphological imperfection were dependent on the activation of cell integrity pathway.
On th e other hand, the sugar structure of cell wall was investigated. As a result, glucan contents decreased to a half in comparison with that of the wild-type and chitin contents increased to about 1.5 times. Remarkable decrease of alpha-glucan was observed in alkaline soluble fraction of the mutant fungi. Alfa-glucan synthase and the encoding gene were not identified, but one candidate protein of which ortholog in A. fumigatus was reported as alpha-glucan synthesis related had Lys-Arg site in N-terminal region. It was presumed that the maturation of the protein by processing should not occur because of the deletion of kexin in A.oryzae. The increment of chitin contents in the alkaline soluble fraction of cell wall was observed, and the enzymatic activity of chitin synthase was risen around 2 times of that of the wild type. Kexin deletion mutant of A.oryzae should not be able to synthesis alpha-glucan, and the perturbation of the cell wall was occurred. As a result of it, cell integrity pathway should be activated and the cell-wall synthesis related enzyme genes should be over-translated.

Research Products

• [Journal Article] Genome sequencing and analysis of Aspergillus oryzae (2005)
  • Author(s): M Machida, K.Asai, -Y Yamagata, -H Kikuchi(全63名)
  • Journal Title: Nature 438 Pages: 1157-1167
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Genome sequencing and analysis of Aspergillus oryzae (2005)
  • Author(s): M Machida, K Asai, M Sano, T, T Kumagai, G Terai, K Kusumoto, T Arima, O Akita, Y Kashiwagi, K Abe, K Gomi, H Horiuchi, K Kitamoto, T Kobayashi, M Takeuchi, D W Denning, J E Galagan, W C Nierman, J Yu, D B.Archer, J W Bennett, D Bhatnagar, T E.Cleveland, N D.Fedorova, O Gotoh, H Horikawa, A Hosoyama, M Ichinomiya, R Igarashi, K Iwashita, P R Juvvadi, M Kato, Y Kato, T Kin, A Kokubun, H Maeda, N Maeyama, J Maruyama, H Nagasaki, T Nakajima, K Oda, K Okada, I Paulsen, K Sakamoto, T Sawano, M Takahashi, K Takase, Y Terabayashi, J R Wortman, O Yamada, Y Yamagata, H Anazawa, Y Hata, Y Koide, T Komori, Y Koyama, T Minetoki, S Suharnan, A Tanaka, K Isono, S Kuhara, N Ogasawara, H Kikuchi
  • Journal Title: Nature 438 Pages: 1157-1161
  • Description: 「研究成果報告書概要(欧文)」より
• [Journal Article] Disorderd cell integrity signaling caused by disruption of the kexB gene in Aspergillus oryzae (2004)
  • Author(s): Miautani O, Nojima A, Yamamoto M, Furukawa K, Fujioka T, Yamagata Y, Abe K, Nakajima T
  • Journal Title: Eukaryotic Cell 3 Pages: 1036-1048
  • Description: 「研究成果報告書概要(和文)」より
• [Journal Article] Disordered cell integrity signaling caused by disruption of the kexB gene in Aspergillus oryzae (2004)
  • Author(s): Mizutani 0., Nojima A., Yamamo M., Furukawa K., Fujioka T., Yamagata Y., Abe K., Nakajima T.
  • Journal Title: Eukaryotic Cell 3 Pages: 1036-1048
  • Description: 「研究成果報告書概要(欧文)」より