Figure 7

The E50K mutant causes reduced uptake of transferrin. (A) HeLa cells were transfected with GFP- tagged wild type optineurin or its mutants, serum starved and incubated with Alexa-594 conjugated transferrin for 20 min. Cells were then fixed and observed with a confocal microscope. (B) The uptake of labelled transferrin was quantitated by measuring the fluorescence intensity in expressing (filled bar) and non-expressing (empty bar) cells. (C) Western blot showing expression of TfR in cells infected with adenoviruses expressing wild-type or E50K mutant optineurin or control viruses. Western blot shows levels of TfR, optineurin and tubulin. (D) HeLa cells were transfected with GFP-tagged wild-type or E50K mutant optineurin. TfR present at the plasma membrane was visualized by staining these cells without permeablilzation using TfR antibody.