Pitstop 2 Is a Potent Inhibitor of Clathrin-Independent Endocytosis
Figure 4
Pitstop 2 inhibits MHCI uptake in other cells lines.
(A) BEAS-2B cells were preincubated with DMSO or 20 µM pitstop 2. Cells were then incubated with Alexa-594 Tfn and antibodies to MHCI in the presence of DMSO or the drugs for 30 min and then processed as described in Figure 2. Lower panel represents the quantification of the integrated intensity of internalized proteins (Tfn and MHCI). The total integrated intensity of each cell was quantified as described in Materials and Methods. The mean fluorescence intensity, as arbitrary units, is plotted with standard deviation for 50 cells in both control and pitstop-treated cells. Both Tfn and MHCI internalization were inhibited at 20 µM pitstop 2 (p<0.001). (B) COS-7 cells were allowed to internalize Alexa-488 Tfn and antibodies to MHCI in the absence or presence of pitstop 2, and quanitified as described in (A). Statistical analysis done for 50 cells with p<0.01 for Tfn inhibition and p<0.001 for MHCI inhibition. The images and quantification shown are from one experiment and similar to that observed in 2 additional experiments. Bars, 10 µm.