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Fig. 6. Effect of AGAP2 on transferrin recycling. (A) AGAP2 promoted transferrin recycling. HeLa cells were transfected with FLAG-AGAP2 for 24 hours. The cells were starved for 30 minutes and then incubated with Rhodamine-conjugated transferrin for 10 minutes (a,b) or 30 minutes (c,d). (B) Effect of [S22N]Rab4 and [S25N]Rab11 on transferrin uptake. Cells were transfected with GFP-[S22N]Rab4 (a,b) and GFP-[S25N]Rab11 (c,d) for 24 hours. Cells were incubated with transferrin for 10 minutes before washing and fixing. (C,D) Interaction of [S22N]Rab4 and [S25N]Rab11 with AGAP2 on transferrin recycling. AGAP2 was transfected into HeLa cells with either GFP-tagged [S22N]Rab4 (C, a-c) or [S25N]Rab11 (D, a-c) for 24 hours. The cells were then incubated with transferrin for 10 minutes. Cells overexpressing AGAP2 were identified by staining for the FLAG tag (arrows). (E) Quantification of intracellular transferrin fluorescence intensity. Intracellular fluorescence intensity of transferrin from single cells was quantified using LSM510 software. The fluorescence intensity of the transfected cells was compared to that of the non-transfected cells on the same coverslip. *P<0.05; **P<0.01 compared with control as analyzed by one-way ANOVA with Tukey post-test.





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