DNA Methylation Changes Of Cucumber In Response To Glucose And Heat And Expression Of Several Differently Methylated Genes

Heat, weak light, dehydration and low temperature and so on are common adverse factors in vegetable cultivation process, which cause the reduction of productivity seriously. Cucumber is one of main cultivated vegetables in our country, when cultivated in open field and greenhouse cultivation usually constrained by these adverse factors. Hence, it is of great significance to investigate the impaction of adversity stress on plants and to explore the way to alleviate stresses. This paper reported the mechanism of dehydration and heat stress mitigated by exogenous glucose. We investigated the exogenous glucose effect on the plant growth parameters and antioxidant enzymes activities of dehydration and heat stressed cucumbers and the exogenous glucose effect on the leaves ultrastructure and the changes of genomic DNA methylation of heat stressed cucumbers. Then the DNA methylation different fragments were separated, cloned, sequenced, sequence analyzed and further analyzed by real time PCR. At the same time, to investigate the role of the exogenous glucose we select osmotic adjustment factor mannitol as control. This paper reveals the mechanism of exogenous glucose alleviate the cucumber dehydration and heat stress in the macro, micro, physiological and molecular levels, which provide a basis for exogenous glucose cultivation of cucumbers and provide references for the study of mechanisms of other exogenous substances to alleviate plant stresses. In this study, the paper use real time fluorescent quantitative PCR to discuss the expression of four differential methylation genes in cucumber leaves, which are under high temperature induced by exogenous glucose, under the weak light induced by hydrogen peroxide, under high temperature induced by paraquat and under low temperature stress. This study provide a theoretical basis for the application of selected genes in cucumber resistance genetic engineering and lay the foundation for elucidating the mechanism of cucumber in response to abiotic stress. The findings are as follows:(1) Effects of exogenous glucose alleviate dehydration and heat stress on cucumber seedling growth parameters, antioxidant enzymes, endogenous glucose, endogenous fructose, SAI, NI and related indexes of antioxidant enzymesIn this study, dehydration and heat stress significantly depressed the growth of (Cucumis sativus L) cv. Jinchun no.4and led to the growth indexes decreased obviously, while exogenous glucose treatment before dehydration or heat stress significantly increased the growth indexes in cucumber seedlings. Therefore, dehydration and heat stress inhibits the growth of cucumber seedlings, while exogenous glucose treatment alleviates the growth inhibition.Compared to control, dehydration and heat stress both increased levels of MDA and ROS (endogenous H2O2and O2-) in the second cucumber leaves in this paper. However, after pretreatment with glucose, the activities of antioxidant enzyme including SOD, CAT, GSH-Px, APX, DHAR, MDHAR and GR and the activities of antioxidants (AsA and GSH) and proline, soluble sugars, SAI and NI were significantly increased. Hence, the levels of MDA and ROS in cucumber leaves were decreased. We concluded that exogenous glucose pretreatment increases antioxidant enzymes and non-enzymatic antioxidants activities in cucumber leaves and decreases the ROS levels, thus reducing the damage of dehydration or heat stress on leaf membrane system.(2) Effects of exogenous glucose on DNA methylation of cucumber genome under heat stressThe changes of DNA methylation in cucumber leaves were measured by MSAP, among these sequences had five MSAP fragments had identity with cucumber genomic sequences. MSAP-2and MSAP-5had identity with coding sequences of gene Csa024621and Csa024512in cucumbers, respectively.(3) The expression of several differently methylated genes in different treatmentsUnder high temperature induced by exogenous glucose, in comparison to control group, heat treatment significantly decreased the transcript levels of gene Csa024621and Csa024512in leaves. Meanwhile, the expression levels of gene Csa024621were significantly decreased and the expression levels of gene Csa024512were significantly increased in leaves of the Glucose+heat treatment when compared to the Heat treatment. Under the weak light induced by hydrogen peroxide, in comparison to control group, low light treatment significantly decreased the transcript levels of gene Csa020125and Csa011787in leaves and the expression levels were significantly increased in leaves of the H2O2+low light group when compared to the low light group. Under high temperature induced by paraquat, compared to control, heat treatment significantly increased the transcript levels of gene Csa025347. In comparison to the heat treatment, treatment of PQ+heat significantly increased the transcript levels of gene Csa025347. Under low tempaterture stress, the expression level of gene Csa017825and Csa010600were significantly decreased in the treatments of MCL and JYL when compared to the MCN and JYN treatment group.