Sulphonation of small molecules by cytosolic sulphotransferases in mammals is an important process in which endogenous molecules are modified for inactivation/activation of their biological effects. Plants possess large numbers of sulphotransferase genes, but their biological functions are largely unknown. Here, we present a functional analysis of the Arabidopsis sulphotransferase AtSOT12 (At2g03760). AtSOT12 gene expression is strongly induced by salt, and osmotic stress and hormone treatments. The T-DNA knockout mutant sot12 exhibited hypersensitivity to NaCl and ABA in seed germination, and to salicylic acid (SA) in seedling growth. In vitro enzyme activity assay revealed that AtSOT12 sulphonates SA, and endogenous SA levels suggested that sulphonation of SA positively regulates SA production. Upon challenging with the pathogen Pseudomonas syringae, sot12 mutant and AtSOT12 overexpressing lines accumulate less and more SA, respectively, when compared with wild type. Consistent with the changes in SA levels, the sot12 mutant was more susceptible, while AtSOT12 over-expressing plants are more resistant to pathogen infection. Moreover, pathogen-induced PR gene expression in systemic leaves was significantly enhanced in AtSOT12 over-expressing plants. The role of sulphonation of SA in SA production, mobile signalling and acquired systemic resistance is discussed.