FIG. 6.

GPI-anchored proteins are transported to the cell surface in immortalized mouse embryonic fibroblasts derived from the cav1-KO mice. (A) Coverslip-grown wild-type (wild-type) and cav1-KO MEFs were infected with a PLAP-expressing adenovirus. Ten hours later, cells were fixed and PLAP and caveolin-1 were visualized by immunofluorescence in unpermeabilized (PLAP) or saponin-permeabilized (caveolin-1) cells. Nuclei were stained with 4′,6′-diamidino-2-phenylindole (DAPI). (B) Wild-type and cav1-KO MEFs were infected with PLAP-expressing adenovirus for 16 h. Cells were pulsed with [³⁵S]methionine for 15 min and chased for 60 min. Surface-exposed PLAP was biotinylated and quantified as described in Materials and Methods. The data shown come from three independent experiments (C) Wild-type and cav1-KO MEFs were grown on coverslips and fixed, and surface-exposed GPI-anchored proteins (GPI-APs) were labeled using an Alexa 546-conjugated aerolysin mutant (Alexa-546-ASSP) on unpermeabilized cells. Total GPI-anchored protein signal was visualized in paraformaldehyde-fixed, saponin-permeabilized MEFs.