ジャーナル: Acta Neuropathol Commun / 年: 2021 タイトル: Correlative light and electron microscopy suggests that mutant huntingtin dysregulates the endolysosomal pathway in presymptomatic Huntington's disease. 著者: Ya Zhou / Thomas R Peskett / Christian Landles / John B Warner / Kirupa Sathasivam / Edward J Smith / Shu Chen / Ronald Wetzel / Hilal A Lashuel / Gillian P Bates / Helen R Saibil / 要旨: Huntington's disease (HD) is a late onset, inherited neurodegenerative disorder for which early pathogenic events remain poorly understood. Here we show that mutant exon 1 HTT proteins are recruited ...Huntington's disease (HD) is a late onset, inherited neurodegenerative disorder for which early pathogenic events remain poorly understood. Here we show that mutant exon 1 HTT proteins are recruited to a subset of cytoplasmic aggregates in the cell bodies of neurons in brain sections from presymptomatic HD, but not wild-type, mice. This occurred in a disease stage and polyglutamine-length dependent manner. We successfully adapted a high-resolution correlative light and electron microscopy methodology, originally developed for mammalian and yeast cells, to allow us to correlate light microscopy and electron microscopy images on the same brain section within an accuracy of 100 nm. Using this approach, we identified these recruitment sites as single membrane bound, vesicle-rich endolysosomal organelles, specifically as (1) multivesicular bodies (MVBs), or amphisomes and (2) autolysosomes or residual bodies. The organelles were often found in close-proximity to phagophore-like structures. Immunogold labeling localized mutant HTT to non-fibrillar, electron lucent structures within the lumen of these organelles. In presymptomatic HD, the recruitment organelles were predominantly MVBs/amphisomes, whereas in late-stage HD, there were more autolysosomes or residual bodies. Electron tomograms indicated the fusion of small vesicles with the vacuole within the lumen, suggesting that MVBs develop into residual bodies. We found that markers of MVB-related exocytosis were depleted in presymptomatic mice and throughout the disease course. This suggests that endolysosomal homeostasis has moved away from exocytosis toward lysosome fusion and degradation, in response to the need to clear the chronically aggregating mutant HTT protein, and that this occurs at an early stage in HD pathogenesis.
材質: Lowicryl HM20 詳細: High pressure frozen samples were freeze substituted in a Leica AFS
詳細
Brain slices were high pressure frozen and freeze substituted
加圧凍結法
装置: OTHER 詳細: The value given for _emd_high_pressure_freezing.instrument is HPM100. This is not in a list of allowed values {'EMS-002 RAPID IMMERSION FREEZER', 'OTHER', 'LEICA EM PACT', 'LEICA EM HPM100', ...詳細: The value given for _emd_high_pressure_freezing.instrument is HPM100. This is not in a list of allowed values {'EMS-002 RAPID IMMERSION FREEZER', 'OTHER', 'LEICA EM PACT', 'LEICA EM HPM100', 'LEICA EM PACT2', 'BAL-TEC HPM 010'} so OTHER is written into the XML file.
Cryo protectant
BSA
切片作成
ウルトラミクロトーム - 装置: Leica EM UC7 / ウルトラミクロトーム - 温度: 20 K / ウルトラミクロトーム - 最終 厚さ: 300 nm
位置合わせマーカー
Manufacturer: EMS / 直径: 10 nm
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電子顕微鏡法
顕微鏡
FEI POLARA 300
撮影
フィルム・検出器のモデル: GATAN K2 SUMMIT (4k x 4k) 平均電子線量: 19.38 e/Å2