Our previous work has indicated that an ethylene-responsive aquaporin gene, Rh-PIP2;1, played an important role in the epidermal cell expansion of rose petals. In this work, we isolated an 896 bp promoter sequence of the Rh-PIP2;1 and found that the promoter was rare in plants, occurring with an Inr motif, but without a TATA box. In transgenic Arabidopsis harboring the Rh-PIP2;1 promoter::GUS construct, the activity of Rh-PIP2;1 promoter was found to be developmental-dependent in almost all of the tested organs, and was particularly active in organs that were rapidly expanding, and in tissues with high water flux capacity. Moreover, the promoter activity was inhibited by ACC, ABA, NaCl, and cold in the roots of 3 or 6-day-old plants, and was increased by GA(3) and mannitol in the rosettes of 9 or 12-day-old plants. Deleting the fragment from -886 to -828 resulted in nearly complete disappearance of the promoter activity in roots, and a substantial decrease in the leaves, hypocotyls and floral organs. Taken together, our results indicated that the Rh-PIP2;1 promoter responded to hormones and abiotic stresses in a developmental- and spatial-dependent manner, and the -886 to -828 region was crucial for the activity of the Rh-PIP2;1 promoter.