WRKY proteins are a large family of transcriptional regulators involved in a variety of biological processes in plants. Here we report functional characterization of a rice WRKY gene, OsWRKY89. RNA gel blot analysis indicated that OsWRKY89 was strongly induced by treatments of methyl jasmonate and UV-B radiation. The transient expression analysis of the OsWRKY89-eGFP reporter in onion epidermal cells revealed that OsWRKY89 was targeted to nuclei. Transcriptional activity assays of OsWRKY89 and its mutants fused with a GAL4 DNA binding domain indicated that the 67 C-terminal amino acids were required for the transcriptional activation and that the leucine zipper region at the N-terminus enhanced its transcriptional activity. Overexpression of OsWRKY89 led to growth retardation at the early stage and reduction of internode length. Scanning electron microscopy revealed an increase in wax deposition on leaf surfaces of the OsWRKY89 overexpression lines and a decrease in wax loading in the RNAi-mediated OsWRKY89 suppression lines. Moreover, extractable and cell-wall-bound phenolic compounds were decreased in the overexpressor lines, but its SA levels were increased. Lignin staining showed an increase in lignification in culms of the overexpressor lines. Interestingly, overexpression of the OsWRKY89 gene enhanced resistance to the rice blast fungus and white-backed planthopper as well as tolerance to UV-B irradiation. These results suggest that OsWRKY89 plays an important role in response to biotic and abiotic stresses.