Fig. 5

Basigin facilitates the early steps of CD98 recycling. a The interaction between basigin and flotillin-1, CD98 and flotillin-1, were analyzed by FRET imaging. Two individual dishes of SMMC-7721 cells were transfected with CD147-dsRed1 or Flotillin-1-eGFP as the acceptor and donor samples, respectively. Two separate dishes were co-transfected with both CD147-dsRed1 and flotillin-1-eGFP as the FRET sample. Using the donor and acceptor samples, CoA and CoB were automatically calculated with FRET software (the grey rectangle area was calculated as background). The FRET ratio is shown on the right. b Effects of flotillin-1 knockdown on the membrane localization of basigin and CD98. SMMC-7721 cells were transfected with siRNAs targeting flotillin-1 for 36 h, then the cells were harvested and the membrane localization of basigin and CD98 was analyzed by FACS. The knockdown effect and total protein levels of basigin and CD98 were analyzed by western blotting. c Co-transfection of Arf6Q67L for 36 h caused the distribution of overexpressed eGFP-N1-CD147(N) (GFP-CD147) to the vacuolar membranes in SMMC7721 cells. d Localization of CD98 in SMMC7721 cells co-expressing GFP-CD147 and Arf6Q67L. SMMC7721 cells co-expressing GFP-CD147 and Arf6Q67L for 36 h were fixed, treated with 0.2 % Triton, and labeled with a primary antibody to CD98 followed by Dylight594-conjugated goat-anti-mouse antibodies. Cell c was transfected with GFP-CD147 only, cell d was transfected with GFP-CD147 and Arf6Q67L (identified by vacuolar membranes), and cells a and b were not transfected and served as a control. Bar, 10 μm