FIG. 5.

The membrane-distal region of the p14 CT drives fusion pore formation. (A) QM5 cells cotransfected with pEGFP and the indicated p14 constructs were cocultured with Vero target cells labeled with calcein red-orange. Cells were resuspended, fixed 9 h posttransfection, and analyzed by flow cytometry. The GFP-positive transfected cells were gated, and the percentages of GFP-positive cells that acquired the aqueous calcein red fluor due to pore formation (indicated above the horizontal threshold line) were quantified and are shown relative to the forward scatter (FSC-H). (B) A similar experiment was performed as described above (A), and results are presented as the calcein red fluorescence distribution of the gated GFP-positive cells (black line tracing). The percentage of cells with calcein red fluorescence above the background fluorescence profile of empty-vector-transfected cells (filled gray histogram) was determined by Overton subtraction of the fluorescence profiles of the two cell populations and is indicated. (C) The dot plots in A and Overton subtraction analysis of the fluorescence profiles in B were quantified and are presented as the means ± SD from one of three experiments conducted in triplicate.