Roles Of Dehydrins And CBF1 Transcription Factor In Plant Drought And Freezing Tolerance

Low temperature and drought represent major environmental constraints limiting growth, development and distribution of plants. A variety of genes are induced both by low temperature and dehydration. These gene products can be classified into two groups. The first group includes proteins that probably function in protecting cells from stresses. The second group of gene products contains protein factors that are involved in further regulation of gene expression and signal transduction in stress response. In this paper, the roles of dehydrins and CBF1 transcription factor were studied in plants under low temperature and dehydration condition, and their possible mechanisms in the development of freezing and drought tolerance were also discussed. I. Roles of dehydrins in plant tolerance to freezing and drought.Accumulations of heat-stable proteins and dehydrins induced by dehydration, low temperature, osmotic stress, high salt or ABA treatment were investigated in protocorm-like-bodies (PLBs) ofDendrobium candidum. Moreover, we described the relationship between these accumulations and the development of tolerance to stresses. Our experimental results were shown as followings:1. PLBs were desiccated and cryopreserved by the air-drying method. Under the controlled system, the dehydration linear equation was expressed at y = -0.1208x + 9.9636. The optimal water content before freezing seemed to be at the range of 0.1 g H2O/g DW to 0.5 g H2O/g DW. Changes in soluble sugars, heat-stable proteins and dehydrins during desiccation of PLBs were analyzed. Extensive accumulation of soluble sugars was observed at water content of about 7.2 g H2O/g DW (after 24 h desiccation), and the sugars contents didn't increased further during the following desiccation. The amount of heat-stable proteins increased significantly when water content decreased to 1.0 g H2O/g DW (after approximately 66 h desiccation). Their molecular masses were 34.3 . 28.7, 19.2,17.0 and 13.3 kDa. Results from immunological detection showed that two bands of the heat-stable proteins with respective molecular masses of 28.7 and 34.3 kDa were dehydrins, which appeared when water content dropped to 1.0 g H2O/g DW. Therefore, it seemed that accumulation of dehydrins happened later than that of soluble sugars. Interestingly, exogenous ABA treatment of PLBs before desiccation could also induce the accumulation of soluble sugars, heat-stable proteins and dehydrins.2. Accumulation of 28.7-kDa dehydrin began to be detectable after 3 days of cold acclimation. Then the level of the dehydrin increased and reached to the maximum at the seventh day. However there were little changes in the level of heat-stable proteins. After 2 hr of exposure to -8.2 ~ -8.0C, the relative survival rate of 3 or 5-day-acclimated PLBs increased to 56.3+22.5% 49.9+19.5% respectively, while that of non-acclimated PLBs was 2.6+0.7%. However, freezing tolerance did not seem to be further enhanced when cold acclimation remained to 7 days. It suggested that acclimated PLBs achieved greater freezing tolerance than non-acclimated controls.3. PEG semilethal concentration was 40% and NaCl semilethal concentration was 200 mmol/L in PLBs by the detection of survival rate of PLBs subjected to various concentrations. Heat-stable protein of 17.0 kDa molecular mass was induced to accumulate by PEG treatment, whose level was increased with the rise of PEG concentration. However, NaCl treatment did not affect the level of heat-stable proteins. Using western blot analysis, it revealed that neither PEG nor NaCl treatment could initiate accumulation of dehydrins in PLBs.II. Studies on enhancement of drought and freezing tolerance by gene transfer ofCBF1.1. CBF1 gene was introduced into tobacco (Nicotiana tabacum) by a plant expression vector pBI121-CBFl containing the CaMV35S promoter using Agrobacterium tumefaciens-mediated transformation. By PCR determination of CBF1 and kanamycin selection of transgenic progeny (Tl, T2 and T3), two puretransgenic lines were obtained...