Fig. 7. Effect of the Hrs-binding region in IL-2Rβ on late-endosomal localization. (A) IL-2Rβ expression on the surface of MEFβ, MEFβ_d268-348, MEFβ_d349-410-2 and HRSdβ cells was examined by flow cytometry as described for Fig. 6A. (B) Aliquots (1.25%) of total lysates from the indicated MEF clones (2x10⁶ cells) were immunoblotted with an anti-IL-2Rβ antibody (C-20) or anti-β-actin antibody. IB, immunoblotting. (C) The indicated cells were grown on coverslips, fixed and double-labeled with an anti-IL-2Rβ antibody (C-20) and an anti-LAMP1 monoclonal antibody or anti-Hrs monoclonal antibody. HRSdβ cells were transfected with GFP-SARA-FYVE, an early endosome marker, and then fixed and labeled with an anti-IL-2Rβ antibody (C20). Red, green and yellow areas indicate IL-2Rβ staining, Hrs or GFP-SARA-FYVE staining and colocalization of the red and green staining, respectively. Scale bars: 5 µm.