The Antioxidant Cyclic 3-Hydroxymelatonin Promotes the Growth and Flowering of Arabidopsis thaliana

Abstract

In plants, melatonin is metabolized into several compounds, including the potent antioxidant cyclic 3-hydroxymelatonin (3-OHM). Melatonin 3-hydroxylase (M3H), a member of the 2-oxo-glutarate-dependent enzyme family, is responsible for 3-OHM biosynthesis. Although rice M3H has been cloned, its roles are unclear, and no homologs in other plant species have been characterized. Here, we cloned and characterized Arabidopsis thaliana M3H (AtM3H). The purified recombinant AtM3H exhibited Km and Vmax values of 100 μM and 20.7 nmol/min/mg protein, respectively. M3H was localized to the cytoplasm, and its expression peaked at night. Based on a 1,1-diphenyl-2-picrylhydrazyl (DPPH) assay, 3-OHM exhibited 15-fold higher antioxidant activity than melatonin. An Arabidopsis M3H knockout mutant (m3h) produced less 3-OHM than the wildtype (WT), thus reducing antioxidant activity and biomass and delaying flowering. These defects were caused by reduced expression of FLOWERING LOCUS T (FT) and gibberellin-related genes, which are responsible for flowering and growth. Exogenous 3-OHM, but not exogenous melatonin, induced FT expression. The peak of M3H expression at night matched the FT expression pattern. The WT and m3h exhibited similar responses to salt stress and pathogens. Collectively, our findings indicate that 3-OHM promotes growth and flowering in Arabidopsis.

Genetic and evolutionary dissection of melatonin response signaling facilitates the regulation of plant growth and stress responses

“…21 While M3H knockout mutant had no difference with WT in salt tolerance. 306 For plant growth, the M2H RNAi rice exhibited reduced germination rate, retarded growth without erect leaf, and delayed leaf senescence, while the ectopic overexpressing rice M2H in tobacco showed dwarfism, leaf necrosis, and early senescence. 307 M2H knockout Arabidopsis mutant showed 60% seed germination at 65h, while M2H overexpressing seed reached 100% germination at 45 h. 308 M3H gene expressed abundantly in rice shoots and roots implying its important role in seedling growth.…”

“…GhM2H149 showed increased cold tolerance of cotton and GhM2H155 only exhibited sensitive to heat stress 21 . While M3H knockout mutant had no difference with WT in salt tolerance 306 . For plant growth, the M2H RNAi rice exhibited reduced germination rate, retarded growth without erect leaf, and delayed leaf senescence, while the ectopic overexpressing rice M2H in tobacco showed dwarfism, leaf necrosis, and early senescence 307 .…”

“…309,310 Arabidopsis M3H knockout mutant exhibited retarded growth with rosette leaves and delayed flowering. 306 Phylogenetic analysis is important for evaluating functional conservation and diversification of gene family (Figure 8B). The large amount of M2H demonstrated that it has undergone significant amplification of gene family during evolution.…”

“…310 The close relationship of M3H from cotton, grape, and soybean with that from Arabidopsis suggested their conservation function in seedling growth and flowering. 306 M3H in cereal plant had an important role in grain yield. 309 Given the rare studies for the functions of M2H and M3H, many questions remain such as the role of M2H and M3H in eudicots; the functions of M2H in biotic stress resistance and plant productive growth including yield and flowering; the functional expansion of M3H in regulating plant abiotic and biotic stress tolerance; the functional overlapping and diversity of M2H and M3H gene family members; the correlation between the expression profile of M2H/M3H and functional diversity.…”

“…149 The expression level of M3H had a significant effect on Arabidopsis flowering by decreasing the expression of GA biosynthetic genes (KS, GA3ox1, and GA3ox2), GA responsive factors (MYB33), and flower inducer (FT). 306 GA coupled with decreased expression of M2H and ASDAC improving rice seed quality with enriched melatonin levels. 313 Under stress condition, melatonin or its metabolites directly or indirectly scavenged ROS and RNS to maintain cellular ROS/RNS balance and then activate stress defense response, which demonstrated that scavenging excessive RNS is a potential mechanism for M2H or M3H in regulating plant life.…”