Figure 9. The domains formed by the Vps subcomplex of the retromer, but not HRS domains, coalesce like WASH domains.

(A) Stable 3T3 cells expressing GFP-Vps35 were treated as in Fig. 6A, then processed for immunofluorescence using WASH antibody and observed by epifluorescence microscopy followed by deconvolution (single planes). Scale bar: 10 µm. (B) corresponds to the insets displayed in (A). Scale bar: 1 µm. Vps35 colocalizes with WASH and behaves similarly to WASH upon actin depolymerization. (C) 3T3 cells were transfected with GFP-Rab5Q79L and DsRed-HRS, and treated as above. Cells were fixed and observed by spinning disk confocal microscopy followed by deconvolution (single planes). Scale bar: 10 µm (1 µm in insets). HRS localizes to discrete domains at the surface of enlarged endosomes, whether or not actin polymerization is prevented.