The Reorientation of T-Cell Polarity and Inhibition of Immunological Synapse Formation by CD46 Involves Its Recruitment to Lipid Rafts
Figure 3
CD46 ligation reduces raft accumulation at the immunological synapse, dependent upon the transmembrane cysteine. (a) and (b), Laurdan-labeled T cells (CD8+ human T cells (a), or CD46Cyt1 transfected MD45 cells (b)) were conjugated to beads coated with antibodies against Tfr or CD3/CD28 as indicated and activated at 37°C for 7 min, fixed and imaged. Where indicated, CD46 was first ligated for 30 min. The GP value of the membranes adjacent to the beads was measured. Each symbol represents one activation site; mean values for each condition are indicated by horizontal bars. Representative images from the CD46 transduced populations in (b) are shown in (c). Laurdan-labeled MD45 cells expressing either CD46Cyt1 (i)–(iv) or CD46Cyt1.C-A (v)–(viii) were untreated ((i), (ii), (v), (vi)) or incubated with CD46-ligating antibodies ((iii), (iv), (vii), (viii)), fixed and imaged. Panels (i), (iii), (v), and (vii) show transmission images with overlaid CD46 staining in red. Panels (ii), (iv), (vi), and (viii) show GP images with the same pseudo-coloring as in Figure 2. The GP values of the membranes adjacent to the beads in the cells shown are 0.273 (ii), 0.181 (iv), 0.374 (vi), and 0.432 (viii). Arrows in (iv) and (viii) show membrane domains that contain CD46Cyt1 and CD46Cyt1.C-A, respectively. The data is representative of 3 experiments. GP values at the site of TCR stimulation were significantly () different between CD46-ligated and not on the TCR-stimulated primary T cells (a), and on the CD46Cyt1-transduced MD45 cells (b).
