{"_buckets": {"deposit": "0c64a0e1-63d0-414a-a015-7f1f03d47146"}, "_deposit": {"created_by": 1, "id": "1346", "owners": [1], "pid": {"revision_id": 0, "type": "depid", "value": "1346"}, "status": "published"}, "_oai": {"id": "oai:ir.soken.ac.jp:00001346", "sets": ["27"]}, "author_link": ["9657", "9658", "9659"], "item_1_biblio_info_21": {"attribute_name": "書誌情報（ソート用）", "attribute_value_mlt": [{"bibliographicIssueDates": {"bibliographicIssueDate": "2001-03-23", "bibliographicIssueDateType": "Issued"}, "bibliographic_titles": [{}]}]}, "item_1_creator_2": {"attribute_name": "著者名", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "桐浴, 隆嘉"}], "nameIdentifiers": [{"nameIdentifier": "9657", "nameIdentifierScheme": "WEKO"}]}]}, "item_1_creator_3": {"attribute_name": "フリガナ", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "キリサコ, タカヨシ"}], "nameIdentifiers": [{"nameIdentifier": "9658", "nameIdentifierScheme": "WEKO"}]}]}, "item_1_date_granted_11": {"attribute_name": "学位授与年月日", "attribute_value_mlt": [{"subitem_dategranted": "2001-03-23"}]}, "item_1_degree_grantor_5": {"attribute_name": "学位授与機関", "attribute_value_mlt": [{"subitem_degreegrantor": [{"subitem_degreegrantor_name": "総合研究大学院大学"}]}]}, "item_1_degree_name_6": {"attribute_name": "学位名", "attribute_value_mlt": [{"subitem_degreename": "博士（理学）"}]}, "item_1_description_1": {"attribute_name": "ID", "attribute_value_mlt": [{"subitem_description": "2001042", "subitem_description_type": "Other"}]}, "item_1_description_12": {"attribute_name": "要旨", "attribute_value_mlt": [{"subitem_description": "Autophagy is the nonselective bulk protein degradation system in the lytic organelle, the lysosome/vacuole, and an example of non-classical vesicular transport from the cytoplasm to the compartment via double-membrane vesicles, autophagosomes.  So far, 15 APG genes essential for autophagy have been cloned in the yeast Saccharomyces cerevisiae, and some APG gene products have been characterised.  However, the molecular mechanism of membrane dynamics of autophagy, in particular the formation of autophagosomes, is still unclear. The WIGS gene encodes a small hydrophilic protein with about 15 kD in mass.  The gene product, Apg8p, drastically increases in abundance by induction of autophagy.  The elevation of amount of Apg8p is regulated transcriptionally.  In spite of the hydrophilic nature, Apg8p is mainly bound to membrane both growth and starvation conditions.\u003cbr /\u003e  Many homologs of Apg8p exist in various eukatyotic organisms.  One of the homologs, LC3, binds to microtubule in vitro. Thus, I examined requirement of microtubule for autophagy morphologically and biochemically using a microtubule-depolymerizing drug, and clarified that microtubule is not necessary for autophagy.\u003cbr /\u003e  After confirming that Apg8p functions in the formation of autophagosome, intracellular localization of Apg8p was assessed by immunofluorescence and immunoelectron microscopic analyses.  The analyses revealed that Apg8p resides on the membrane structures at the various stages in autophagosome formation, autophagosomes and autophagic bodies. Apg8p is the first identified molecule that traces the whole process of membrane dynamics in autophagy.  Immunoelectron microscopy also revealed that Apg8p is more enriched on the membranes of the intermediate structures of autophagosome than on those of mature autophagosomes.  The result indicated that Apg8p should directly function at the step of autophagosome formation and suggested that it may be detached from the membranes before autophagosome is formed up completely.  Furthermore, the observation of the formation process of autophagosomes traced with Apg8p provided a new proposal that autophagosomal membrane may be generated de novo but not from pre-existing membrane cistema.\u003cbr /\u003e  As described above, Apg8p seemed to be a key molecule for elucidating the molecular mechanism of autophagosome formation. Thus, I performed further characterization of Apg8p and discovered that Apg8p undergoes a novel reversible modification.  Apg8p is produced as the precursor form in cells.  The newly synthesized Apg8p is cleaved by a novel cysteine protease, Apg4p, to remove the original carboxy-terminal arginine residue, and thereby a Gly residue becomes the carboxy-terminal residue of the protein.  Subsequently, Apg8p forms a covalent conjugate with a general glycerophospholipid, phosphatidylethanolamine (PH), which is a major component of biological membrane. Apg8p binds to PE via amide bond between carboxyl group of the carboxy-terminal glycine of Apg8p and amino group of PE.  The formation of the adduct, designated Apg8p-PE, requires Apg7p and Apg3p.  Finally, Apg8p-PE is deconjugated to Apg8p and PH by cleavage action of Apg4p. Apg7p and Apg3p were already identified as the modification enzymes for Apg8p, which are equivalent to E1 and E2 enzymes for ubiquitination, respectively.   Furthermore, the mode of action of Apg4p, which cleaves both newly synthesized Apg8p and Apg8p-PE, well resembles that of some deubiquitinating enzymes which cleave both the precursor of ubiquitin and the adducts of ubiquitin with substrate.  The overall similarity demonstrates that ubiquitination-like system involves in the formation of Apg8p-PB.  So far, there had been no report that ubiquitination and the related systems are utilized to protein- lipidation.  This is the first evidence that ubiquitination-like modification mediates protein-lipidation.  At the point of protein-lipidation, this study showed the first example that protein is covalently conjugated to phosphatidylethanolamine.\u003cbr /\u003e  The formation and deconjugation of Apg8p-PE is essential for formation of autophagosomes.  Free Apg8p is originally loosely membrane bound or soluble.  The reversible modification changes the membrane-binding state of Apg8p.  Apg8p turns to be bound to membrane tightly by the conjugation with PE, and it is returned to loosely-membrane bound or soluble by the deconjugation.  These features are consistent with morphological evidence that Apg8p is well localized along the membrane of autophagosome intermediates but little detected on mature autophagosome.  The regulation of the membrane-binding state of Apg8p by the reversible modification is essential for autophagosome formation.", "subitem_description_type": "Other"}]}, "item_1_description_7": {"attribute_name": "学位記番号", "attribute_value_mlt": [{"subitem_description": "総研大甲第537号", "subitem_description_type": "Other"}]}, "item_1_select_14": {"attribute_name": "所蔵", "attribute_value_mlt": [{"subitem_select_item": "有"}]}, "item_1_select_8": {"attribute_name": "研究科", "attribute_value_mlt": [{"subitem_select_item": "生命科学研究科"}]}, "item_1_select_9": {"attribute_name": "専攻", "attribute_value_mlt": [{"subitem_select_item": "X2 分子生物機構論専攻"}]}, "item_1_text_10": {"attribute_name": "学位授与年度", "attribute_value_mlt": [{"subitem_text_value": "2000"}]}, "item_1_text_20": {"attribute_name": "業務メモ", "attribute_value_mlt": [{"subitem_text_value": "（2018年2月16日）本籍など個人情報の記載がある旧要旨・審査要旨を個人情報のない新しいものに差し替えた。承諾書等未確認。要確認該当項目修正のこと。"}]}, "item_creator": {"attribute_name": "著者", "attribute_type": "creator", "attribute_value_mlt": [{"creatorNames": [{"creatorName": "KIRISAKO, Takayoshi", "creatorNameLang": "en"}], "nameIdentifiers": [{"nameIdentifier": "9659", "nameIdentifierScheme": "WEKO"}]}]}, "item_files": {"attribute_name": "ファイル情報", "attribute_type": "file", "attribute_value_mlt": [{"accessrole": "open_date", "date": [{"dateType": "Available", "dateValue": "2016-02-17"}], "displaytype": "simple", "download_preview_message": "", "file_order": 0, "filename": "甲537_要旨.pdf", "filesize": [{"value": "315.7 kB"}], "format": "application/pdf", "future_date_message": "", "is_thumbnail": false, "licensetype": "license_11", "mimetype": "application/pdf", "size": 315700.0, "url": {"label": "要旨・審査要旨 / Abstract, Screening Result", "url": "https://ir.soken.ac.jp/record/1346/files/甲537_要旨.pdf"}, "version_id": "9f9b9678-673d-4171-b2aa-156fb82c7404"}]}, "item_language": {"attribute_name": "言語", "attribute_value_mlt": [{"subitem_language": "eng"}]}, "item_resource_type": {"attribute_name": "資源タイプ", "attribute_value_mlt": [{"resourcetype": "thesis", "resourceuri": "http://purl.org/coar/resource_type/c_46ec"}]}, "item_title": "New Insight in the Membrane Dynamics of Autophagy by Studies on Apg8p", "item_titles": {"attribute_name": "タイトル", "attribute_value_mlt": [{"subitem_title": "New Insight in the Membrane Dynamics of Autophagy by Studies on Apg8p"}, {"subitem_title": "New Insight in the Membrane Dynamics of Autophagy by Studies on Apg8p", "subitem_title_language": "en"}]}, "item_type_id": "1", "owner": "1", "path": ["27"], "permalink_uri": "https://ir.soken.ac.jp/records/1346", "pubdate": {"attribute_name": "公開日", "attribute_value": "2010-02-22"}, "publish_date": "2010-02-22", "publish_status": "0", "recid": "1346", "relation": {}, "relation_version_is_last": true, "title": ["New Insight in the Membrane Dynamics of Autophagy by Studies on Apg8p"], "weko_shared_id": 1}