Ultrastructural characterization of the delimiting membranes of isolated autophagosomes and amphisomes by freeze-fracture electron microscopy.
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/microm2, whereas isolated lysosomes had about 2000 particles/microm2). 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/microm2), 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.
Fengsrud
M
M
Department of Cell Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Montebello, Oslo.
Erichsen
E S
ES
Berg
T O
TO
Raiborg
C
C
Seglen
P O
PO
D000818:Animals; D001343:Autophagy / Q000502:physiology*; D001710:Biotin / Q000378:metabolism; D020285:Cryoelectron Microscopy / Q000379:methods; D005614:Freeze Fracturing; D007425:Intracellular Membranes / Q000378:metabolism / Q000648:ultrastructure; D008565:Membrane Proteins; D010588:Phagosomes / Q000648:ultrastructure*; D051381:Rats; D014617:Vacuoles / Q000648:ultrastructure
D008565:Membrane Proteins; D001710:Biotin