Fig. 2.
Vesicles containing engineered Eqt fuse with the plasma membrane. (A) Blockage of export from the Golgi results in retention of Eqt-SM. HeLa cells expressing Eqt-SM (tagged with oxGFP) were incubated at 20 °C for 2 h (top row) to arrest export from the TGN and then transferred to 37 °C for 30 min. The arrows point to cytoplasmic puncta containing Eqt-SM that appeared after release of the 20 °C Golgi export block. Maximum projections of a z series are shown. (Scale bar: 10 μm.) (B) Representative TIRFM frames for Eqt-SM and Eqt-sol. (C) TIRFM showing vesicles containing Eqt-SM and Eqt-sol fused with the plasma membrane. The green and red traces indicate the average normalized fluorescence intensities for Eqt-SM (n = 413) and Eqt-sol (n = 268) vesicle fusion events, respectively (prefusion intensity defined as 0; postfusion intensity defined as 1). SDs for each point are shown. (D) Rates of Eqt-SM and Eqt-sol exocytic events as determined by TIRFM imaging and expressed as the number of fusion events per area (μm2) per time (min). SEMs are indicated. The rates are not statistically different (P ≤ 0.06).