Fig. 4

Knockdown of EHD1 suppresses β2AR endocytic recycling. a, b Representative images of the colocalization of EHD1 (red) and β2AR (green) in A549 (a) and NCI-H1650 (b) cells. c A549 and NCI-H1650 cell lysates were immunoprecipitated with the anti-EHD1 antibody, and the immunoprecipitates were subjected to Western blot analysis. d A549 and NCI-H1650 cell lysates were immunoprecipitated with the anti-β2AR antibody, and the immunoprecipitates were subjected to Western blot analysis. e, f Representative images and colocalization of β2AR and Rab11 in A549 and A549/Sh cells (e) and in A549/Sh/Ctrl and A549/Sh/R cells (f). β2AR was visualized using an anti-β2AR antibody (green fluorescence), Rab11 was visualized using an anti-Rab11 antibody (red fluorescence), and their colocalization is shown in the right-hand panels (merge; yellow fluorescence). Scale bars, 10 μm. g, h EHD1 stabilizes β2AR. A549 cells were incubated in serum-free 1640 medium for 16 h and then treated with ISO (10 μm) for the indicated times in the presence of cycloheximide (CHX, 20 μg/ml). The cell lysates were analyzed by Western blot