In this study we tested the hypothesis that loss of TBC1D5 function will enhance the activity of the retromer complex by increasing the levels of membrane-associated retromer CSC proteins necessary for sorting cargo into tubules. We show that the loss of TBC1D5 expression upon RNA interference does indeed increase the levels of endosomally localised VPS26 and VPS35 proteins. As these proteins are components of the retromer CSC, which is a hub for recruiting other machinery to endosomes, we found that increasing the endosomal retromer CSC by TBC1D5 knockdown also enhances the association of the retromer CSC with accessory proteins such as Fam21 (and other components of the WASH complex, e.g. strumpellin and KIAA1033), Rme-8 (also known as DNAJC13), Snx3 and Snx27. In fact, loss of TBC1D5 expression can rescue the recruitment of Fam21 to the endosome in cells expressing the PD-causing VPS35 D620N mutant. The interaction of Fam21 with the retromer CSC requires the binding of VPS29 to VPS35, and TBC1D5 interacts with VPS29 (Helfer et al., 2013; Harbour et al., 2010). Thus, it is possible that the gain in
