Fig 8.

Us2 colocalizes predominantly with Rab11, a marker of recycling endosomes. (A to C) Vero cells were cotransfected with plasmids encoding HSV-2 Us2 and either Rab5-GFP (A), Rab7-GFP (B), or Rab11-GFP (C). Plasmids encoding Rab5-, Rab7-, and Rab11-GFP fusion proteins were a kind gift from C. Parish, Cornell University. Cells were fixed at 24 h posttransfection and were stained for Us2. Images of stained cells were captured by confocal microscopy. Rab signals are EGFP fluorescence and are displayed in green. The Us2 signal is red. Nuclei were detected using the DNA stain Hoechst 33342 (blue). Arrowheads in panel C highlight the colocalization of Us2 and Rab11 signals. Representative images are shown. (D) Quantification of the colocalization of Us2-labeled puncta with Rab5, Rab7, and Rab11. The data are presented as percentages of Us2-containing puncta that are also positive for Rab5, Rab7, or Rab11. A total of 12 fields of cells and 249 Us2 puncta were analyzed. The analysis was performed in a manner similar to that for the quantification of Us2-transferrin colocalization, for which results are shown in Fig. 7D and E. Error bars represent the standard errors of the means observed for differences between different fields of cells for each Rab protein analyzed.