Fig. 6. GRIP domain overexpression affects Golgi to plasma membrane transport. HeLa cells were transiently transfected with expression vectors for VSV-G-EGFP (2.5 µg/six-well dish) and either vector alone (a-c'), C312(Y2187A) (d-f') or C312 (g-m') at 5µg/six-well dish. Cells were grown at 39°C for 1 day and then fixed either before (a,d,g) or after shifting to 32°C for the indicated times. Cells were analyzed by fluorescence microscopy after immunostaining with anti-HA and RRX-conjugated secondary antibodies (' panels); EGFP was visualized directly (non-' panels). (g-k') Examples of cells with similar anti-HA staining intensities to those shown in d-f'; (l,l') an example of a cell with low C312 expression, and (m,m') an example of a cell with very high C312 expression. All ' panels were taken at the same exposure time on samples prepared and analyzed on the same day. The `spotty' appearance of VSV-G-EGFP localization in a, d and g may result from a fixation artifact.

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