Molecular Mechanism Of Ethylene Regualtion Of Freezing Tolerance In Arabidopsis

In many parts of the world, low temperature stress adversely affects plant growth and development. After short period of exposure to cold treatment above0℃, plants grown in temperate regions enhance tolerance to freezing stress, this process is called’cold acclimation’. Significant progress towards better understanding the molecular basis of cold tolerance in plants has been made in recent years. Up to date, the ICE1-CBF-COR transcriptional regulatory cascade has been extensively studied. Cold stress can rapidly activate the important transcription factors CBFs (C-repeat factors), which in turn bind to the C-repeat (CRT)/dehydration-responsive element of cold responsive (COR) genes, and confer the freezing tolerance of plants. The CBF genes are directly regulated by the master regulator ICE1(Inducer of CBF expression1). Moreover, The ICE1protein has been found to be regulated by several translational modifications.The phytohormone ethylene regulates multiple aspects of plant growth and development processes. Recent studies have shown that ethylene is involved in regulation of plant responses to many abiotic stresses. Cold stress alters endogenous ethylene levels in many plant species, but the role of ethylene in freezing tolerance is unclear. In this study, we found that ethylene negatively regulates plant responses to freezing stress in Arabidopsis.Freezing tolerance was decreased in wild-type Col and the ethylene overproducing mutant etol seedlings when treated with the ethylene precursor1-aminocyclopropane-1-carboxylic acid (ACC), whereas an inhibitor of ethylene biosynthesis aminoethoxyvinyl glycine (AVG) increased the freezing tolerance of both wild-type and etol plants. Furthermore, the ethylene overproducing mutants etol, eto2and eto3showed reduced freezing tolerance compared to the wild type. By contrast, the hexuple and heptuple mutants of ACC synthases responsible for ethylene biosynthesis displayed enhanced freezing tolerance. In addition, ethylene biosynthesis was dramatically inhibited in wild type and etol mutant by cold stress. These results suggest a negative role of ethylene on plant cold tolerance.The freezing tolerance of wild-type plants was increased by the ethylene perception antagonist Ag". etrl-1and ein4-1are gain-of-function of ethylene receptors, and they showed enhanced freezing tolerance compared to the wild type. EIN2and EIN3are key positive regulators in the ethylene signaling. The loss-of-function mutant ein2-5, ein3-1and ein3eill were more tolerant to freezing stress than the wild-type plants. By contrast, the constitutive ethylene response mutant ctrl-1and EIN3-overexpressing plants exhibited reduced freezing tolerance. Further genetic and biochemical analyses revealed that EIN3negatively regulates the expression of CBFs by specifically binding to the conserved EBS (EIN3binding sites) motif in the promoter regiones. Besides, cold stress stabilized EIN3protein in an EIN2-dependent manner, which is possibly caused by the degradation of the F-box protein EBF1.Thus, our study demonstrates that ethylene negatively regulates cold signaling through direct repressing the transcription of CBFs by EIN3in Arabidopsis. These findings provide a new perspective on the genetic control of freezing tolerance and our understanding of the molecular basis of the effects of ethylene on plant responses to adverse environments, as well as the crosstalk of ethylene and cold signaling.