Heterotrimeric G-proteins play an important role in plant signalling pathways. The plant hormone methyl jasmonate (MeJA) can induce stomatal closure in many plant species. The signal cascade in MeJA-induced stomatal closure has been studied previously. However, the function of G proteins in this process has not yet been evaluated. In this study, the stomatal movement induced by MeJA in the wild-type Arabidopsis thaliana (L. Heynh.) (WS), Gα subunit loss-of-function mutant gpa1-1 and gpa1-2 guard cells were measured. Further, the transmembrane ion flux (H+, Ca2+ and K+) and reactive oxygen species (ROS) experiments were performed in guard cells from WS, GDP-β-S pre-treated WS, gpa1-1 and gpa1-2 using non-invasive micro-test technique (NMT) and confocal technique. It was observed that the MeJA-induced stomatal closure was abolished in guard cells of gpa1 mutants. GDP-β-S pre-treatment and gpa1 mutants impaired the MeJA-activated H+ efflux, Ca2+ influx and K+ efflux. The accumulation of ROS in gpa1-1 and gpa1-2 guard cells was also lower than that in WS guard cells under MeJA treatment. These results suggested that Gα subunits are involved in regulating the signal events in JA signal pathway and stomatal closure.