PC1 is completely encapsulated in large COPII-coated vesicles. (A) Two-color 3D STORM revealed PC1 (cyan) residing inside of large SEC31A (magenta) cages in KI6 cells after 7.5 h of induced KLHL12 overexpression. (i) Overview of an area of the cell imaged by STORM. Small (80- to 100-nm-diameter) canonical SEC31A puncta did not colocalize with PC1 (0%, n = 76 puncta, n = 8 experiments). Examples in dashed inset are magnified in ii as merged image, in iii as SEC31A, and in iv as PC1. Large SEC31A puncta of diameter larger than 300 nm colocalized with PC1 (76.3%, n = 36 puncta, n = 8 experiments). An example in solid inset is magnified in v as merged image, in vi as SEC31A, and in vii as PC1. A virtual cross section in the xz plane of the large COPII vesicle is shown in viii as merged image, in ix as SEC31A, and in x as PC1. (B) Triple-color 3D STORM imaging shows endogenous PC1 (cyan) encapsulated by endogenous KLHL12 (yellow) and SEC31A (magenta) in Saos-2 cells grown at steady state. xy maximum projection of the overview is shown in i and of a magnified vesicle in ii–v. Virtual cross sections in the xz plane are shown in x–xii and yz in vi–ix. Merged three-color channel is shown in i, ii, vi, and x; SEC31A in iii, vii, and xi; PC1 in iv, viii, and xii; and KLHL12 in v, ix, and xiii. Bars: (Ai) 1 µm; (Bi) 2 µm; (magnified views) 100 nm.