Characterization of Isoprene Synthase from Arundo donax L. in Relation to Abiotic Stress Tolerance

Isoprene (2-methyl-1,3-butadiene, C5H8) is a five carbon volatile hydrocarbon compound, which is synthesized through methylerythritol phosphate (MEP) pathway in the chloroplasts of many plant species. Isoprene has been suggested to enhance plants tolerance to stressful conditions because its emission could be stimulated when leaves are subjected to or recovering from environmental stresses. Physiological function(s) of isoprene emission has been studied for a long time, and previous studies well documented that isoprene emission can enhance plant thermotolerance and oxidative stress resistance; however, whether isoprene emission may increase plant tolerance to drought stress is less studied, and the molecular mechanisms of isoprene emitting plants in response to drought-related stress have never been investigated before. In this study, we characterized the physiological function of the first IspS gene (AdoIspS) from a monocot species, Arundo donax L., in response to different abiotic stresses. First of all, Phenotypic differences under normal growth condition between AdoIspS transgenic and Col-0 wild type plants were observed as follows: earlier flowering time, tendency to higher leaf biomass even though no significant difference, and a higher seed production derived from higher numbers of branching in AdoIspS transgenic plants. The results suggested that isoprene emission may provide a selective advantage to plants by enhancing plant growth rate. In order to investigate the protective function of isoprene emission in response to abiotic stresses, first, the enhanced thermotolerance was tested by analyzing the survival ability between AdoIspS transgenic plants and Col-0. The results indicate that after recovering from the heat shock, AdoIspS transgenic plants exhibited a higher survival rate, chlorophyll content and fresh weight at different developmental stages, which revealed that the transformation of AdoIspS gene isolated from a monocot species can provide similar thermotolerance to isoprene non-emitter as IspSs from dicots. Second, we investigated AdoIspS transgenic plants sensitivity to exogenous abscisic acid (ABA). AdoIspS transgenic plants showed a decreased sensitivity to exogenous ABA application at both germination and seedling stages. qPCR results indicate that under the treatment of ABA on root part, the regulation of ABA-induced genes and ABA biosynthesis genes in root is not affected by the presence of AdoIspS; however, in AdoIspS transgenic leaves, ABA-response mark genes are up-regulated and ABA biosynthesis genes are down-regulated, which supports the idea that isoprene emission may reduce ABA biosynthesis and accumulation in leaves. Therefore, the growth inhibition from exogenous ABA treatment decreased in AdoIspS transgenic plants. To elucidate whether and how the dehydration tolerance was altered or not in later developmental stage of AdoIspS transgenic plants, AdoIspS transgenic Arabidopsis lines were further applied in dehydration stress studies. The results of water loss test and stomatal aperture assay consistently demonstrated that isoprene-emitting plants may have a reduced requirement for ABA to tolerant dehydration stress. Additionally, the survival rate, lipid peroxidation test and dehydration-related gene expression were measured to analyze AdoIspS transgenic plant tolerance to dehydration stress at the whole-plant level, and the results indicate that the better dehydration tolerance displayed in AdoIspS transgenic lines maybe due to the reduced ROS accumulation in isoprene-emitting leaves. These findings suggested that AdoIspS gene plays an important role in plant's tolerance to abiotic stress conditions. Understanding the regulatory mechanisms of AdoIspS gene in response to abiotic stresses could help us find out why isoprene-emitting plants cope better with transient stress events in nature.