Reactive oxygen species (ROS), including HO, contribute to oxidative stress and may cause cancer initiation and progression. However, at low concentrations, HO can regulate signaling pathways modulating cell growth, differentiation, and migration. A few mammalian aquaporins (AQPs) facilitate HO diffusion across membranes and participate in tumorigenesis. AQP3 and AQP5 are strongly expressed in cancer tissues and AQP3-mediated HO transport has been related to breast cancer cell migration, but studies with human AQP5 are lacking. Here, we report that, in addition to its established water permeation capacity, human AQP5 facilitates transmembrane HO diffusion and modulates cell growth of AQP5-transformed yeast cells in response to oxidative stress. Mutagenesis studies revealed that residue His173 located in the selective filter is crucial for AQP5 permeability, and interactions with phosphorylated Ser183 may regulate permeation through pore blockage. Moreover, in human pancreatic cancer cells, the measured AQP5-mediated HO influx rate indicates the presence of a highly efficient peroxiporin activity. Cell migration was similarly suppressed by AQP3 or AQP5 gene silencing and could be recovered by external oxidative stimuli. Altogether, these results unveiled a major role for AQP5 in dynamic fine-tuning of the intracellular HO concentration, and consequently in activating signaling networks related to cell survival and cancer progression, highlighting AQP5 as a promising drug target for cancer therapies.