Functional Analysis Of A Wheat Expansin Gene TaEXPA2

Wheat is cultivated as one of the most significant food crops worldwide, and drought stress is an acute environmental condition that can severely block the growth and productivity of wheat. Through genetic engineering to improve the adversity of wheat adaptability is the development direction of wheat breeding. Related gene function research can lay theoretical foundation for the wheat genetic engineering breeding, genetic resources reserves.Expansins are cell wall proteins that are grouped into two main families, α-expansins and β-expansins, and they are implicated in the control of cell extension via the disruption of hydrogen bonds between cellulose and matrix glucans. In 1989, Cosgrove was first isolated the expansin from cucumber hypocotyls. There is increasing evidence for the involvement of expansins in the plant development process and the responses of various plant species to various stresses. However, due to large wheat genome, genetic background is complex, and conversion efficiency is low, wheat protein gene function research reported less extension.Expansins are encoded by multi- gene families. There exist similarities and differences in gene structure, protein structure, biochemical characteristics, evolutionary relationships, as well as other aspects between family members. The EXPA and EXPB are the two biggest gene family, research the most clearly. O ur previous study indicated that wheat expansin expression responds to abiotic stress. The ectopic expression or the root-specific expression of Ta EXPB23, a wheat β-expansin gene, improved the drought tolerance of transgenic tobacco. In this study, to discover the similarities and d ifferences in the functions of α- and β-expansins, we selected a wheat α-expansin gene, Ta EXPA2(Gen Bank ID: AAT94292.1), and investigated its involvement in plant drought tolerance and development.First,we cloned the promoter region of Ta EXPA2 and analyzed its cis-elements. Then,we analyzed expression patterns of Ta EXPA2 in wheat and studied its functions in drought and salt tolerance in transgenic tobacco. Third, we explored the relations between Ta EXPA2 and the cell wall POD enzymes by Ta EXPA2-overexpressing Arabidopsis and a mutant atexpa2. The main results are listed as follws:(1) In this study, we isolated the promoter fragment of Ta EXPA2 and predicted the putative cis-regulatory elements with Plant CARE. There are several types of cis-regulatory elements in the promoter fragment of Ta EXPA2, includingcis-acting elements involved in light responses, phytohormone signaling, and abiotic stress responsiveness, as well as an enhancer- like element involved in anoxic specific inducibility meristem expression, cell cycle regulation, zein metabolism regulation, seed-specific regulation, circadian control, and so on.(2) Through transient expression of 35S::Ta EXPA2-GFP in onion epidermal cells, Ta EXPA2 was localized to the cell wall.(3) We performed q RT-PCR analysis to characterize the expression patterns of Ta EXPA2. The transcript levels were highest in leaf. It strongly increased after 12 h of PEG treatment compared to the control.(4) Constitutive expression of Ta EXPA2 in tobacco improved seed production by increasing capsule number, not seed size, without having any effect on plant growth patterns.(5) The transgenic tobacco exhibited a significantly greater tolerance to water-deficiency stress than did wild-type(WT) plants. We found that under drought stress, the transgenic plants maintained a better water status. The accumulated content of osmotic adjustment substances, such as proline, in Ta EXPA2 transgenic plants was greater than that in WT plants. Transgenic plants also displayed greater antioxidative competence as indicated by their lower malondialdehyde(MDA) content, relative electrical conductivity, and reactive oxygen species(ROS) accumulation than did WT plants. This result suggests that the transgenic plants suffer less damage from ROS under drought conditions. The activities of some antioxidant enzymes as well as expression levels of several genes encoding key antioxidant enzymes were higher in the transgenic plants than in the WT plants under drought stress.(6) Our results suggest that Ta EXPA2 overexpression confers salt stress tolerance to the transgenic plants, which may be associated with the homeostasis of Na+/K+, improved water status and antioxidant competence under salt stress. When leaf discs of wild type(WT) plants were incubated in cell wall protein extracts from the leaves of transgenic tobacco plants, their chlorophyll content was higher under salt stress. By contrast, the increased salt tolerance of transgenic tobacco plants was inhibited when the leaf discs were treated with antibody against Ta EXPA2 protein.(7) We studied the involvement of abscisic acid(ABA) signaling in the response of Ta EXPA2 to salt stress in wheat. The accumulation of Ta EXPA2 protein in wheat leaves was induced by Na Cl and ABA treatments, but inhibited by FLU( fluridone, an ABA inhibitor). When Na Cl and FLU were administered to the wheat seedlings simultaneously, the Na Cl- induced increase of Ta EXPA2 protein was inhibited, suggesting that ABA signaling was involved in the Na Cl- induced increase of Ta EXPA2 protein. However, the expression levels of tobacco ABA biosynthesis genes were lower in Ta EXPA2-overexpressing tobacco plants than in WT plants under both normal and salt stress conditions, although their expression was upregulated by Na Cl treatment.(8) Using homologous gene in Arabidopsis mutants atexpa2 studied the physiological and molecular mechanisms of overexpression Ta EXPA2 improved oxidation resistance of transgenic plants. The expression of Ta EXPA2 can induced by H2O2 treatment. Overexpressing Ta EXPA2 enhanced the tolerance to oxidative stress in transgenic Arabidopsis, and the activity of cell wall POD. The decreased oxidative stress tolerance and activity of cell wall POD in atexpa2 were further confirmed the relation between expansin and cell wall POD. Research of transgenic Arabidopsis antioxidant capacity increase of physiological mechanism, found that the transgenic arabidopsis antioxidant capacity increase with plant cell wall is close to the key of peroxidase. The results for explaining expansin pro tein how to enhance stress tolerance of plants is of great significance.