Fig. 2.

EDEM1 vesicles are formed outside of the transitional ER. (A–C) Confocal immunofluorescence, HepG2 hepatoma cells, staining for EDEM1 (A), calnexin (B), and overlay (C, codistribution is indicated by different shades of yellow). EDEM1 staining is essentially punctate along with some elongated structures, whereas that for calnexin is reticular (B). (C) EDEM1 and calnexin staining partially overlaps in elongated structures. (D and E) Ultrathin frozen sections from HepG2 cells with immunogold labeling for EDEM1 in the ER lumen (arrowheads) and ER-associated smooth vesicles (arrows). Clathrin-coated vesicles (cv in D) are unlabeled. (F–I) Immunoperoxidase labeling for EDEM1 reveals staining in the lumen of rough ER (F, arrowhead), in ER buds (G), and vesicles pinching-off the ER (H, arrowhead) or close to the ER (I, arrowheads). (K) Four serial sections from a HepG2 cell with luminal EDEM1 labeling (black dots) in parts of ER cisternae. Arrowhead in K2 points to ER membrane-associated EDEM1 staining and arrow to cytoplasmic EDEM1 staining. cp in K 1–3: clathrin-coated pit. (Scale bars, 10 μm in A–C; 60 nm in D and E; 80 nm in F; 95 nm in G; 155 nm in H; 130 nm in I; and 400 nm in K.)