The vacuolar H+-ATPase (V-ATPase) was first identified as an electrogenic proton pump that acidifies the lumen of intra- and extracellular compartments. The acidic pH generated by V-ATPase is important for a wide range of cellular processes as well as acidification-independent processes such as secretion and membrane fusion. In addition to these housekeeping functions, recent studies implicate V-ATPase in the direct regulation and function of signaling pathways. In this review, we describe recent findings on the functions of V-ATPase in growth regulation and tissue physiology. •V-ATPase-regulated endocytic trafficking is essential for signal transduction.•V-ATPase functions as an intrinsic part of various signaling mechanisms.•Catalytic activity of V-ATPase provides feedback control for intracellular signaling.