2-Hydroxymelatonin, Rather Than Melatonin, Is Responsible for RBOH-Dependent Reactive Oxygen Species Production Leading to Premature Senescence in Plants

Abstract

Unlike animals, plants amply convert melatonin into 2-hydroxymelatonin (2-OHM) and cyclic 3-hydroxymelatonin (3-OHM) through the action of melatonin 2-hydroxylase (M2H) and melatonin 3-hydroxylase (M3H), respectively. Thus, the effects of exogenous melatonin treatment in plants may be caused by melatonin, 2-OHM, or 3-OHM, or some combination of these compounds. Indeed, studies of melatonin’s effects on reactive oxygen species (ROS) production have reported conflicting results. In this study, we demonstrated that 2-OHM treatment induced ROS production, whereas melatonin did not. ROS production from 2-OHM treatment occurred in old arabidopsis leaves in darkness, consistent with an ethylene-mediated senescence mechanism. Transgenic tobacco plants containing overexpressed rice M2H exhibited dwarfism and leaf necrosis of the upper leaves and early senescence of the lower leaves. We also demonstrated that 2-OHM-mediated ROS production is respiratory burst NADPH oxidase (RBOH)-dependent and that 2-OHM-induced senescence genes require ethylene and the abscisic acid (ABA) signaling pathway in arabidopsis. In contrast to melatonin, 2-OHM treatment induced senescence symptoms such as leaf chlorosis and increased ion leakage in arabidopsis. Senescence induction is known to begin with decreased levels of proteins involved in chloroplast maintenance, including Lhcb1 and ClpR1. Together, these results show that 2-OHM acts as a senescence-inducing factor by inducing ROS production in plants.