Recycling of AQP2 occurs through a temperature- and bafilomycin-sensitive trans-Golgi-associated compartment

The exo- and endocytotic pathway in which aquaporin-2 (AQP2) travels between the plasma membrane and intracellular vesicles is only partially characterized. It is known that the antidiuretic hormone vasopressin induces a translocation of AQP2 from an intracellular to a plasma membrane location, both in kidney collecting duct principal cells and in transfected epithelial cells. Here we provide evidence suggesting that while AQP2 shifts from an intracellular location to the cell surface in response to vasopressin, AQP2 also constitutively recycles through a similar pathway in transfected LLC-PK(1) cells even in the absence of hormonal stimulation. Incubating cells at 20 degrees C blocks AQP2 recycling in a perinuclear compartment, regardless of whether vasopressin is present. The H(+)-ATPase inhibitor bafilomycin A1 also blocks the recycling pathway of AQP2 in a perinuclear compartment adjacent to the Golgi in the presence and absence of vasopressin stimulation, indicating a role of vesicle acidification in both the constitutive and regulated recycling of AQP2. Colocalization of AQP2 with clathrin, but not with giantin, after both bafilomycin treatment and a 20 degrees C block suggests that the compartment in which recycling AQP2 is blocked may be the trans-Golgi, and not cis- and medial-Golgi cisternae.