Functional Analysis Of Annexin Gene (AnnSp2), Enhances Abiotic Stress Tolerance In Transgenic Tomato

Drought and high salinity are two major abiotic stresses that significantly affect agricultural crop productivity worldwide.Annexins are multigene family that plays an essential role in plant stress responses and various cellular processes.Here,the AnnSp2 gene was cloned from drought-resistant wild tomato(Solanum pennellii)and functionally characterized in cultivated tomato.AnnSp2 protein was localized in the nucleus and had higher expression in leaves,flower and fruit.It was induced by several phytohormones and some abiotic stresses.Tomato plants overexpressing AnnSp2 had increased tolerance to drought and salt stress,as determined by analysis of various physiological parameters.AnnSp2-transgenic plants were less sensitive to ABA during the seed germination and seedling stages.However,under drought stress,the ABA content significantly increased in the AnnSp2-overexpressing plants,inducing stomatal closure and reducing water loss,which underlay the plants' enhanced stress tolerance.Furthermore,scavenging reactive oxygen species(R)S),higher total chlorophyll content,lower lipid peroxidation levels,increased peroxidase activities(including APX,CAT and SOD)and higher levels of proline were observed in AnnSp2-overexpressing plants.These results indicate that overexpression of AnnSp2 in transgenic tomato improves salt and drought tolerance through ABA synthesis and the elimination of ROS.The main results are as follows:1.AnnSp2 was cloned and sequenced,which encoded an annexin protein of 316 amino acids.Expression profile analysis using RT-PCR showed that AnnSp2 transcript was abundant in leaves and flowers,AnnSp2 protein was predominantly localized in the nucleus.The gene was strongly induced by several phytohormones and some abiotic stresses such as dehydration,salinity and hormone like abcisic acid(ABA).2.Overexpression vectors of AnnSp2 in tomato,containing the target gene via Agrobacterium-mediated transformation,and a total of 16 transgenic plants were obtained.Abiotic stress tolerance assays were applied on three AnnSp2-overexpressing lines.Tomato plants overexpressing AnnSp2 had increased tolerance to drought and salt stress,as determined through physiological analysis of the germination rate,root growth,survival rate,leaf water loss and Chl content.3.AnnSp2-transgenic plants were less sensitive to ABA during the seed germination and seedling stages that underlay the plants' enhanced stress tolerance.Moreover,scavenging reactive oxygen species(ROS),higher total chlorophyll content,lower lipid peroxidation levels,increased peroxidase activities(including APX,CAT and SOD)and higher levels of proline were observed in AnnSp2-overexpressing plants.4.Results from both germination,seedling experiments,by altering expression of stress related genes indicate that overexpression of AnnSp2 in transgenic tomato improves salt and drought tolerance through ABA synthesis and the elimination of ROS.