Ultrastructural characterization of the delimiting membranes of isolated autophagosomes and amphisomes by freeze-fracture electron microscopy
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- 作者：Monica Fengsrud ; Egil S. Erichsen ; Trond O. Berg ; Camilla Raiborg ; Per O. Seglen
- 关键词：Rat ; hepatocytes ; freeze-fracture ; epon ; electron microscopy ; autophagy ; autophagosome ; amphisome ; lysosome ; transmembrane protein ; intramembrane particle
- 刊名：European Journal of Cell Biology
- 出版年：2000
- 出版时间：December 2000
- 年：2000
- 卷：79
- 期：12
- 页码：871-882
- 全文大小：317 K
文摘
The delimiting membranes of isolated autophagosomes from rat liver had extremely few transmembrane proteins, as indicated by the paucity of intramembrane particles in freeze-fracture images (about 20 particles/μm2, whereas isolated lysosomes had about 2000 particles/μm2). The autophagosomes also appeared to lack peripheral surface membrane proteins, since attempts to surface-biotinylate intact autophagosomes only yielded biotinylation of proteins from contaminating damaged mitochondria. All the membrane layers of multilamellar autophagosomes were equally particle-poor; the same was true of the autophagosome-forming, sequestering membrane complexes (phagophores). Isolated amphisomes (vacuoles formed by fusion between autophagosomes and endosomes) had more intramembrane particles than the autophagosomes (about 90 particles/μm2), and freeze-fracture images of these organelles frequently showed particle-rich endosomes fusing with particle-poor or particle-free autophagosomes. The appearence of multiple particle clusters suggested that a single autophagic vacuole could undergo multiple fusions with endosomes. Only the outermost membrane of bi- or multilamellar autophagic vacuoles appeared to engage in such fusions.