Table II.
Effects of Microinjected GTP-γ-S or IgGs Directed Against Erd2p, β-COP, or p23 on the Retrograde Transport of CTX-A–K63 from the Golgi to the ER in Vero Cells
| Localization of CTX-A–K63 | ||||
|---|---|---|---|---|
| Golgi | ER | |||
| % of counted microinjected cells | ||||
| Controls | 5 | 95 | ||
| BSA | 6 | 94 | ||
| GTP-γ-S | 90 | 10 | ||
| Controls | 5 | 95 | ||
| Preimmune IgGs | 6 | 94 | ||
| Anti-Erd2p | 58 | 42 | ||
| Controls | 2 | 98 | ||
| Preimmune IgGs | 7 | 93 | ||
| Anti–β-COP IgGs | 74 | 26 | ||
| Controls | 4 | 96 | ||
| Preimmune IgGs | 9 | 91 | ||
| Anti–p23 IgGs | 72 | 28 | ||
Uptake of CTX–K63 (0.5 μg/ml) was initiated as in Fig. 2. After 15–20 min the cells were microinjected and the transport of CTX-A–K63 followed for another 80 min. The cells were then fixed and analyzed for the distribution of CTX-A–K63 by immunofluorescence. Microinjected cells were identified by either dye-labeled BSA mixed with GTP-γ-S, or by mixing Cy2-labeled primary IgGs to the corresponding antibodies at a 1:3 ratio. The effect of microinjection itself was controlled in experiments where preimmune IgGs were injected. In each group 25–30 microinjected cells were analyzed.