FIG 6.

Cellular internalization and localization of the sortilin S15D mutant mimicking constitutive PAK phosphorylation. (A) Cellular internalization. HEK cells stably expressing the wild-type or an S15D mutated sortilin receptor were incubated for 1 h on ice with monoclonal mouse antisortilin clone F11 antibodies. Cells were washed; incubated at 37°C for 0, 5, or 10 min; and finally fixed in 4% paraformaldehyde and permeabilized. Following fixation, cells were stained with Alexa Fluor 488-conjugated donkey anti-mouse antibody (green), and the nuclei were visualized by using Hoechst stain (blue). (B) Immunocytochemistry of sortilin in transfected HEK cells showing the localization of wt sortilin and sortilin S15D. Cells were stained with monoclonal mouse antisortilin clone F11 antibody (green), and nuclei were visualized with Hoechst stain (blue).