Most cartilaginous fishes are principally marine species, and only a limited number of species, including bull shark (Carcharhinus leucas), have the capacity to inhabit both seawater and freshwater environments. To reveal the mechanisms of euryhalinity in cartilaginous fishes, we conducted a transfer experiment of captive bull sharks from seawater to freshwater and performed RNA-sequencing using two seawater- and freshwater-acclimated bull shark kidneys, because the kidney is the only organ that is able to excrete excess water from the body in the hypoosmotic environment. We further conducted differential gene expression analysis to search for genes up- and down-regulated in the freshwater-acclimated bull shark kidneys.