FIGURE 5:

Tfn dynamics in cells depleted of dynactin subunits. (A and B) siRNA-treated Cos7 cells were incubated with Alexa Fluor 546-Tfn for (A) 5 min and (B) 15 min and then fixed and costained for TfR. Scale bar: 5 μm. The insets are 2×. (C) Cells (labeled with Tfn for increasing times) exhibiting colocalization of Tfn with perinuclear TfR were counted (mean ± SD of three independent experiments; n > 300 cells/condition). *, Values significantly different from controls (p < 0.05). (D) Kinetics of Tfn uptake and recycling. Cells were labeled with Tfn for increasing lengths of time (0–60 min). At the time points indicated, cells were fixed, and Tfn fluorescence (50 cells/condition) was quantified. (E) Analysis of particle movement in cells labeled with Alexa Fluor 555-Tfn for 2 min, then imaged in the interval between 10 and 15 min of chase. The data represent particle movements occurring in a total of 800 s of observation (80 s/cell, 10 cells/condition). (F) Cos7 cells cotransfected with control or p27 siRNA, plus pCAGIG-p27 as indicated (rescue). Data were collected as in (E).