INVs are involved in recycling α5β1 integrin. (A) Bubble plot of total intensities from immunoprecipitates prepared from RPE-1 cells expressing GFP or GFP-TPD54 (n_exp = 3). (B) Bubble plot of total spectral counts from BioID2 experiments (n_replicates = 6, n_exp = 2). In A and B, all TPD52-like proteins, Rabs and integrins detected in the dataset are shown. Size of bubbles is normalized to the most abundant protein detected per experiment. (C) ELISA-based quantification of integrin α5 recycling over time in siCtrl (gray line) or siTPD54-treated (blue line) RPE1 cells. **, P < 0.001; *, P < 0.05. Bars show SD. n_exp = 3. (D) Representative confocal micrographs of integrin α5 recycling. RPE-1 cells (siCtrl or siTPD54 treated) were surface labeled using anti-integrin α5 (VC5) and allowed to recycle for the indicated times. The experiment was repeated three times, and similar results were also obtained using an alternative α5 antibody (SNAKA51). Scale bar, 10 µm.