Fig. 3

The subcellular localization of CD28 is altered in the presence of Nef or Vpu. CD4⁺ Sup-T1 cells were infected with VSV-G pseudotyped NL4.3 viruses encoding or lacking Nef and/or Vpu. Infected cells were stained for CD28 and the lysosomal marker LAMP-1 and visualized by widefield microscopy. a Shown are representative infected (GFP⁺) cells (left) and a graphical representation of CD28 (blue) and LAMP-1 (red) fluorescence intensity relative to the maximum along the illustrated line (right). The scale bar indicates 10 μm and the vertical lines labelled PM indicate where the illustrated line meets the plasma membrane (PM). Insets in the merged panel illustrate the GFP channel. b The percentage overlap between CD28 and LAMP-1 is calculated based on the mean (± SD) Manders’ overlap coefficients from at least 30 cells and 3 independent experiments. (UI: uninfected; LAMP-1: Lysosomal-associated membrane protein 1; SD: standard deviation; ****p ≤ 0.0001)