+データを開く
-基本情報
登録情報 | データベース: EMDB / ID: EMD-50068 | |||||||||
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タイトル | Electron tomogram of ER-nuclear envelope junction of HeLa cell in interphase | |||||||||
マップデータ | ||||||||||
試料 |
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キーワード | Endoplasmic reticulum / Nuclear envelope / Organelle contact site / Membrane junction / CELL CYCLE | |||||||||
生物種 | Homo sapiens (ヒト) | |||||||||
手法 | 電子線トモグラフィー法 / クライオ電子顕微鏡法 / ネガティブ染色法 | |||||||||
データ登録者 | Bragulat-Teixidor H / Otsuka S | |||||||||
資金援助 | オーストリア, 2件
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引用 | ジャーナル: EMBO Rep / 年: 2024 タイトル: The endoplasmic reticulum connects to the nucleus by constricted junctions that mature after mitosis. 著者: Helena Bragulat-Teixidor / Keisuke Ishihara / Gréta Martina Szücs / Shotaro Otsuka / 要旨: Junctions between the endoplasmic reticulum (ER) and the outer membrane of the nuclear envelope (NE) physically connect both organelles. These ER-NE junctions are essential for supplying the NE with ...Junctions between the endoplasmic reticulum (ER) and the outer membrane of the nuclear envelope (NE) physically connect both organelles. These ER-NE junctions are essential for supplying the NE with lipids and proteins synthesized in the ER. However, little is known about the structure of these ER-NE junctions. Here, we systematically study the ultrastructure of ER-NE junctions in cryo-fixed mammalian cells staged in anaphase, telophase, and interphase by correlating live cell imaging with three-dimensional electron microscopy. Our results show that ER-NE junctions in interphase cells have a pronounced hourglass shape with a constricted neck of 7-20 nm width. This morphology is significantly distinct from that of junctions within the ER network, and their morphology emerges as early as telophase. The highly constricted ER-NE junctions are seen in several mammalian cell types, but not in budding yeast. We speculate that the unique and highly constricted ER-NE junctions are regulated via novel mechanisms that contribute to ER-to-NE lipid and protein traffic in higher eukaryotes. | |||||||||
履歴 |
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-構造の表示
添付画像 |
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-ダウンロードとリンク
-EMDBアーカイブ
マップデータ | emd_50068.map.gz | 12.8 GB | EMDBマップデータ形式 | |
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ヘッダ (付随情報) | emd-50068-v30.xml emd-50068.xml | 9.5 KB 9.5 KB | 表示 表示 | EMDBヘッダ |
画像 | emd_50068.png | 231.4 KB | ||
Filedesc metadata | emd-50068.cif.gz | 4.1 KB | ||
アーカイブディレクトリ | http://ftp.pdbj.org/pub/emdb/structures/EMD-50068 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-50068 | HTTPS FTP |
-検証レポート
文書・要旨 | emd_50068_validation.pdf.gz | 508.2 KB | 表示 | EMDB検証レポート |
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文書・詳細版 | emd_50068_full_validation.pdf.gz | 507.7 KB | 表示 | |
XML形式データ | emd_50068_validation.xml.gz | 4 KB | 表示 | |
CIF形式データ | emd_50068_validation.cif.gz | 4.5 KB | 表示 | |
アーカイブディレクトリ | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-50068 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-50068 | HTTPS FTP |
-関連構造データ
-リンク
EMDBのページ | EMDB (EBI/PDBe) / EMDataResource |
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-マップ
ファイル | ダウンロード / ファイル: emd_50068.map.gz / 形式: CCP4 / 大きさ: 17.8 GB / タイプ: IMAGE STORED AS SIGNED INTEGER (2 BYTES) | ||||||||||||||||||||
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ボクセルのサイズ | X=Y=Z: 4.51 Å | ||||||||||||||||||||
密度 |
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対称性 | 空間群: 1 | ||||||||||||||||||||
詳細 | EMDB XML:
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-添付データ
-試料の構成要素
-全体 : HeLa Kyoto cell
全体 | 名称: HeLa Kyoto cell |
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要素 |
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-超分子 #1: HeLa Kyoto cell
超分子 | 名称: HeLa Kyoto cell / タイプ: cell / ID: 1 / 親要素: 0 |
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由来(天然) | 生物種: Homo sapiens (ヒト) / 株: HeLa |
-実験情報
-構造解析
手法 | ネガティブ染色法, クライオ電子顕微鏡法 |
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解析 | 電子線トモグラフィー法 |
試料の集合状態 | cell |
-試料調製
緩衝液 | pH: 7.4 詳細: DMEM without Riboflavin and Phenol Red, containing 10% FBS, 1% Pen/Strep, and 50 nM SiR-DNA |
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染色 | タイプ: NEGATIVE / 材質: Uranyl acetate and lead citrate |
糖包埋 | 材質: Agar 100 Epoxy resin 詳細: Frozen cells were substituted in 0.1% uranyl acetate (UA), 2% Osmium tetroxide and 5% H2O in acetone following this temperature ramp: -90 C to -80 C for 10 hours, -80 C to -30 C for 10 hours, ...詳細: Frozen cells were substituted in 0.1% uranyl acetate (UA), 2% Osmium tetroxide and 5% H2O in acetone following this temperature ramp: -90 C to -80 C for 10 hours, -80 C to -30 C for 10 hours, -30 C for 4 hours, -30 C to 0 C for 6 hours, 0 C to 20 C for 4 hours, 20 C for 5-6 hours. Afterwards, samples were washed three times in pure acetone for at least 10 minutes each, and subsequently infiltrated with Agar 100 Epoxy resin. The resin infiltration was done progressively at room temperature with increasing concentrations of resin in acetone (3:1 for 2-3 hours, 1:1 for 2-3 hours, and 1:3 overnight). Infiltration with pure resin was done at room temperature for at least 5 hours. Resin was polymerized at 60 C for 72 hours. |
凍結 | 凍結剤: NITROGEN |
加圧凍結法 | 装置: OTHER 詳細: High pressure freezing chamber was 1 mm thick, 6.0 mm diameter, with central cavities 5.0 mm x 5.0 mm x 25 um deep. The chamber had been in contact with 1-hexadecene.. The value given for _em_ ...詳細: High pressure freezing chamber was 1 mm thick, 6.0 mm diameter, with central cavities 5.0 mm x 5.0 mm x 25 um deep. The chamber had been in contact with 1-hexadecene.. The value given for _em_high_pressure_freezing.instrument is Leica EM ICE. This is not in a list of allowed values {'OTHER', 'LEICA EM PACT', 'LEICA EM HPM100', 'EMS-002 RAPID IMMERSION FREEZER', 'BAL-TEC HPM 010', 'LEICA EM PACT2'} so OTHER is written into the XML file. |
Cryo protectant | 20% Ficoll-PM400 |
切片作成 | ウルトラミクロトーム - 装置: Leica Ultracut UCT / ウルトラミクロトーム - 温度: 298 K / ウルトラミクロトーム - 最終 厚さ: 250 |
位置合わせマーカー | Manufacturer: Cytodiagnostics / 直径: 15 nm |
-電子顕微鏡法
顕微鏡 | FEI TECNAI F20 |
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撮影 | フィルム・検出器のモデル: FEI EAGLE (4k x 4k) / 平均電子線量: 40.0 e/Å2 |
電子線 | 加速電圧: 200 kV / 電子線源: FIELD EMISSION GUN |
電子光学系 | 照射モード: FLOOD BEAM / 撮影モード: BRIGHT FIELD / 最大 デフォーカス(公称値): 0.5 µm / 最小 デフォーカス(公称値): 0.2 µm |
実験機器 | モデル: Tecnai F20 / 画像提供: FEI Company |
-画像解析
最終 再構成 | アルゴリズム: BACK PROJECTION / 使用した粒子像数: 110 |
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