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

The exo- and endocytotic pathway in which aquaporin-2 (AQPB) travels betwee n the plasma membrane and intracellular vesicles is only partially characte rized. It is known that the antidiuretic hormone vasopressin induces a tran slocation of AQP2 from an intracellular to a plasma membrane location, both in kidney collecting duct principal cells and in transfected epithelial ce lls. Here we provide evidence suggesting that while AQPB shifts from an int racellular location to the cell surface in response to vasopressin, AQPB al so constitutively recycles through a similar pathway in transfected LLC-PK1 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 als o blocks the recycling pathway ofAQP2 in a perinuclear compartment adjacent to the Golgi in the presence and absence ofvasopressin stimulation, indica ting a role of vesicle acidification in both the constitutive and regulated recycling ofAQP2. Colocalization of AQP2 with clathrin, but not with giant in, 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.