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2002 Fiscal Year Final Research Report Summary

MOLECULAR-BIOLOGICAL RESEARCH ON VACUOLAR FUNCTION OF FILAMENTOUS FUNGI USING KOJI-MOLDS

Research Project

Project/Area Number 12460041
Research Category

Grant-in-Aid for Scientific Research (B)

Allocation TypeSingle-year Grants
Section一般
Research Field 応用微生物学・応用生物化学
Research InstitutionTHE UNIVERSITY OF TOKYO

Principal Investigator

KITAMOTO Katsuhiko  Graduate School of Agricultural and Life Sciences, Professor, 大学院・農学生命科学研究科, 教授 (20272437)

Project Period (FY) 2000 – 2002
Keywordskoji mold / vacuoles / A. nidulans / EGFP / carboxy peptidase / VPS mutants / A. oryzae
Research Abstract

In yeast S. cerevisiae, vacuoles play very important roles in pH and osmotic regulation, protein degradation and storage of amino acids, small ions as well as polyphosphates. In filamentous fungi, however, little is known about vacuolar functions at a molecular level. Several genes homologous to yeast VPS (VPS1, VPS45, and VPS21) and VAM (VAM4, and VAM6) genes were isolated from Aspergillus nidulans and designated the genes as vpsA, vpsB, vpsC, avaA, and avaB, respectively. These gene disruption experiments of A. nidulans showed poor growth and contained fragmented vacuoles. These results suggested that the genes isolated are involved in vacuolar biogenesis and functions. In addition, cpyA (carboxy peptidase Y homologue) gene was also isolated as a marker enzyme of vacuoles from A. nidulans. The cpyA gene was fused with EGFP (Enhanced Green Fluorescent Protein) gene and the resulting fusion gene was introduced into A. nidulans and A. oryzae, respectively. Vacuoles are clearly visualized by a fluorescent microscope in the living fungi by expressing of CPY-EGFP. Eighteen hfc (hyper EGFP fluoresce in conidia) mutants, 20 dfc (darker fluorescence in conidia) mutants, and 22 hfm (hyper EGFP fluorescence in the medium) mutants were isolated from A. oryzae expressing CPY-EGFP as a parent strain using a FACS ( fluorescence activated cell sorter) or a fluorescent micro plate reader. hfc-1 mutant showed very poor growth in alkaline medium. Fragmented vacuoles, which are similar to class B vps mutants in yeast, were observed in hfc-3, hfc-4, dfc-14, and hfm-4 mutants. Accumulation of small spherical structure near vacuolar membrane was observed in dfc-14, and hfm-4 mutants in alkaline medium, which are similar to class E vps mutants.

  • Research Products

    (12 results)

All Other

All Publications (12 results)

  • [Publications] Y.Tarutani, K.Ohsumi, M.Arioka, H.Nakajima, K.Kitamoto: "Cloning and characterization of Aspergillus nidulans vpsA gene which is involved in vacuolar biogenesis"Gene. 268. 23-30 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] K.Ohsumi, Y.Matsuda, H.Nakajima, K.Kitamoto: "Cloning and characterization of the cpyA gene encoding intracellular carboxypeptidase from Aspergillus nidulans"Biosci. Biotech. Biochem. 65. 1175-1180 (2001)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] M.Ohneda, M.Arioka, H.Nakajima, K.Kitamoto: "Visualization of Vacuoles in Aspergillus oryzae by expression of CPY-EGFP"Fungal Genetics and Biology. 37. 29-38 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] J.Maruyama, H.Nakajima, K.Kitamoto: "Observation of EGFP-visualized nuclei and distribution of vacuoles in Aspergillus oryzae arpA null mutant"FEMS Microbiol Letters. 206. 57-61 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] K.Ohsumi, M.Arioka, H.Nakajima, K.Kitamoto: "Cloning and characterization of a gene (avaA) from Aspergillus nidulans encoding a small GTPase involved in vacuolar biogenesis"Gene. 291. 77-84 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y.Kuroki, P.R.Juvvadi, M.Arioka, H.Nakajima, K.Kitamoto: "Cloning and characterization of vmaA, the gene encoding a 69-kDA catalytic subninit of the vacuolar H^+-ATPase during pH mediated growth of Aspergillus oryzae"FEMS Microbiol Letters. 209. 277-282 (2002)

    • Description
      「研究成果報告書概要(和文)」より
  • [Publications] Y. Tarutani, K. Ohsumi, M. Arioka, H. Nakajima, K. Kitamoto: "Cloning and characterization of Aspergillus nidulans vpsA gene which is involved in vacuolar biogenesis"Gene. 268. 23-30 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] K. Ohsumi, Y. Matsuda, H. Nakajima, K. Kitamoto: "Cloning and characterization of the cpyA gene encoding intracellular carboxypeptidase from Aspergillus nidulans"Biosci. Biotech. Biochem.. 65. 1175-1180 (2001)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] M. Ohneda, M. Arioka, H. Nakajima, and K. Kitamoto: "Visualization of Vacuoles in Aspergillus oryzae by expression of CPY-EGFP"Fungal Genetics and Biology. 37. 29-38 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] J. Maruyama, H. Nakajima, K. Kitamoto: "Observation of EGFP-visualized nuclei and distribution of vacuoles in Aspergillus oryzae arpA null mutant"FEMS Microbiol Letters. 206. 57-61 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] K, Ohsumi, M. Arioka, H. Nakajima, K. Kitamoto: "Cloning and characterization of a gene (avaA) from Aspergillus nidulans encoding a small GTPase involved in vacuolar biogenesis"Gene. 291. 77-84 (2002)

    • Description
      「研究成果報告書概要(欧文)」より
  • [Publications] Y. Kuroki, P.R.Juvvadi, M. Arioka, H. Nakajima, Kitamoto: "Cloning and characterization of vmaA, the gene encoding a 69-kDA catalytic subninit of the vacuolar H^+-ATPase during pH mediated growth of Aspergillus oryzae"FEMS Microbiol Letters. 209. 277-282 (2002)

    • Description
      「研究成果報告書概要(欧文)」より

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Published: 2004-04-14  

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