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Fig. 6. Rab1 is the only Rab essential for ER to cell surface transport. (A,B) VSV G transport assays were performed as described in the Materials and Methods on HeLa cells expressing the wild-type and catalytically inactive arginine finger point mutant TBC-domain proteins indicated. The cell surface appearance of VSV G was measured for all conditions (A). The bar graph (B) shows the inhibition in transport observed with the various wild-type (grey bars), and catalytically inactive (open bars) TBC-domain proteins. (C) VSV G transport assays were performed in HeLa cells expressing Myc-tagged wild-type (WT) or catalytically inactive arginine finger point mutant (R105A) TBC1D20. After 60 minutes of transport cells were fixed and surface stained for VSV G (red), then permeabilized and stained for the Myc epitope (blue). Total VSV G was observed by GFP fluorescence (green) at all time points. (D) VSV G transport assays were performed in HeLa cells treated with control, Rab1, p115, or GM130 siRNA duplexes for 72 hours. After 60 minutes of transport cells were fixed and surface stained for VSV G (red), and total VSV G was observed by GFP fluorescence (green). (E) VSV G transport assays were performed in HeLa cells expressing Myc-tagged wild-type (WT), dominant-negative (N121I) or constitutive active (Q67L) point mutants of Rab1. After 60 minutes of transport the cells were fixed and surface stained for VSV G (red), then permeabilized and stained for the Myc epitope (blue). Total VSV G was observed by GFP fluorescence (green). Bars, 10 µm.
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