Technical Abstract: Endothall, an older commercial herbicide, and cantharidin, a natural product from the blister beetle (Epicauta spp.), are close chemical analogues. A comparison of the effect of endothall and cantharidin on plants revealed a similarity in their level of phytotoxicity on both Arabidopsis thaliana and Lemna paucicostata. Cantharidin is a potent inhibitor of animal serine/threonine protein phosphatases. Protein phosphatases and kinases maintain a sensitive balance between phosphorylated and dephosphorylated forms of proteins playing important roles in signal transduction pathways. In this study, the phytotoxicity of endothall and cantharidin was directly related to their ability to inhibit plant serine/threonine protein phosphatase activity. Both compounds acted as slow irreversible inactivators of the serine/threonine phosphatases activity. Several cellular pathways affected by the inhibition of these protein phosphatases by cantharidin in A. thaliana by transcriptome analyses and endothall caused an identical change in gene expression, but in a more pronounced way. Therefore, the molecular target site of endothall in plants is similar to that of cantharidin in animals, namely, serine/threonine protein phosphatases responsible for regulating an array of biochemical processes. This mode of action is unlike any other commercial herbicide.