Mitogen‐activated protein kinase pathways are required for melatonin‐mediated defense responses in plants

Melatonin enhances pathogen resistance by inducing the expression of a number of plant defense‐related genes. To examine whether the melatonin‐mediated pathogen resistance is associated with mitogen‐activated protein kinase ( MAPK ) cascades, Arabidopsis and tobacco leaves were treated with melatonin and investigated for MAPK activation using an antiphospho‐p44/42 MAPK (Erk1/2) monoclonal antibody. Two MAPK s, MPK 3 and MPK 6, were activated rapidly and transiently by 1  μ m melatonin treatment in Arabidopsis . Its tobacco ortholog MAPK s were also activated. The activation of MPK 3 and MPK 6 by 2‐hydroxymelatonin and N ‐acetylserotonin was also observed, albeit to a lesser degree than that by melatonin. Furthermore, MAPK activation by melatonin was uncoupled from G‐protein signaling, because melatonin efficiently activated two MAPK s in a G‐protein β knockout mutant ( agb1 ). Suppression of both MPK 3 and MPK 6 in transgenic Arabidopsis exhibited significant decreases in the induction of defense‐related gene expression and pathogen resistance relative to wild‐type plants. Using an array of MAP kinase kinase ( MKK ) knockout mutants, we found that four MKK s, namely MKK 4, MKK 5, MKK 7, and MKK 9, are responsible for the activation of MPK 3 and MPK 6 by melatonin, indicating that melatonin‐mediated innate immunity is triggered by MAPK signaling through MKK 4/5/7/9‐ MPK 3/6 cascades.