Fig. 2. Structure and expression of TrkA-Ros and SHP-1 fluorescent fusion proteins. (A) Schematic presentation of fluorescent fusion protein constructs. Ros is used as a TrkA-Ros chimerical protein, which allows stimulation of the kinase by nerve growth factor (NGF). Protein domains and important amino acid residues are indicated. (B) Expression of TrkA-Ros-ECFP and EYFP-SHP1-WT in HEK293 cells. TL, transmitted light; bar, 10 µm. (C) Localization of TrkA-Ros-ECFP in the plasma membrane of HEK293 cells shown by immuno-electron microscopy. Membranes of HEK293 cells expressing TrkA-Ros-ECFP and wild-type SHP-1, or mock-transfected cells for control, were freeze-fractured and TrkA-Ros-ECFP was detected with anti-GFP antibodies, followed by immuno-gold labeled secondary antibody. The inner plasma membrane surface, identified by morphological criteria, is shown. Outer surfaces, nuclear membrane or mitochondria showed no labeling (not depicted).