Mitogen-activated protein kinase (MAPK) cascades serve as unified signaling modules in plant development and defense response. Previous reports demonstrated an essential role of Arabidopsis GLIP1, a member of the GDSL-like-motif lipase family, in both local and systemic resistance. GLIP1 expression is highly induced by pathogen attack. However, the one or more signaling pathways involved are unknown. Here, we report that two pathogen-responsive MAPKs, MPK3 and MPK6, are implicated in regulating gene expression of GLIP1 as well as GLIP3 and GLIP4. After gain-of-function activation, MPK3 and MPK6 can strongly induce the expression of GLIP1, GLIP3, and GLIP4. Both GLIP1 and GLIP3 contribute to the plant resistance to Botrytis cinerea. WRKY33, a MPK3/MPK6 substrate, is essential for the MPK3/MPK6-dependent GLIP1 induction. In addition, WRKY2 and WRKY34, two close homologs of WRKY33, have a minor effect in MPK3/MPK6-regulated GLIP1 expression in B. cinerea-infected plants. Chromatin immunoprecipitation-quantitative polymerase chain reaction analysis demonstrated that the GLIP1 gene is a direct target of WRKY33. In addition, we demonstrated that MPK3/MPK6-induced GLIP1 expression is independent of ethylene and jasmonic acid, two important hormones in plant defense. Our results provide insights into the regulation of the GLIP family at the transcriptional level in plant immunity.