Fig. 4

CD98 is retained in the ER and Rab5 positive endosomes of live cancer cells in the absence of basigin. a Schematic of the different basigin expression constructs and pcDNA3.1-dsRED1-CD98 (dsRED1-CD98). Note that the sequences encoding eGFP or dsRED1 were inserted into the signal peptide and coding sequence of basigin and CD98, respectively. b Representative distribution of overexpressed CD98 (red) in SMMC7721 cells and K7721 cells. SMMC-7721 cells (5 × 10⁴ cells) were cultured on coverslips and transfected with 2 μg of dsRED1-CD98 for 36 h. The distribution of dsRED1-CD98 was visualized with a confocal microscope. The percentage of CD98 (red) blocked cells was determined by scoring > 100 cells. The data are the mean of the “% of dsRED1-CD98 blocked cells” from three independent experiments ± SED. Bar, 10 μm. ** P < 0.01. c Cytoplasmic distribution of overexpressed CD98 (red) in K7721 cells. After the transfection of dsRED1-CD98 as described in part B, the distribution of the endoplasmic reticulum (ER) was revealed with ER-Tracker^TM. Bar, 5 μm. EGFP-Rab5A Q79L or GFP-Rab11 (1 μg) was co-transfected with 1 μg of dsRED1-CD98 to visualize the endosomal distribution of cytoplasmically retained CD98. (Pearson’s coefficient is indicated as numerical data on the right of each panel, n = 3). Bar, 10 μm. d Representative distribution of overexpressed CD98 (red) after co-transfection of different basigin expression constructs. Three different basigin expression constructs (green) were co-transfected with dsRED1-CD98 in K7721 cells for 36 h, and the distribution of overexpressed CD98 (red) was then visualized. Bar, 10 μm