Regulation And Biological Function Of BZR1 Transcription Factor At Low Temperature

As an important environmental factor,low temperature adversely affects the growth,development,metabolism and geographical distribution of plants.In order to adapt to the environment,plants have evolved sophisticated mechanisms to protect against cold stress.In recent years,it has been found that brassinosteroid(BR)is involved in adaptation of plant on low temperature,and coldinduced BR-related transcription factor BZR1 can affect gene expression and improve the freezing tolerance of plant.However,the mechanism of low-temperature regulation of BZR1 transcriptional activity remains unclear,and the biological function of BZR1 at low temperatures remains to be further studied.This study used physiological and biochemical,genetic,and bioinformatics methods to explore the these issues.The main results are as follows.1.Western blot analysis showed that cold treatments induced BZR1 dephosphorylation not only in Col-0,but also in BR biosynthesis or signaling-defective mutants.Moreover,cold treatmemt also induced BZR1 dephosphorylation in Col-0 and bri1-116 with a 35S:BZR1-YFP construct.These results indicate that cold-induced BZR1 dephosphorylation is independent of BR synthesis and perception.2.CHIP-q PCR and q PCR analysis showed that cold stress promotes the binding of BZR1 to its target gene promoters,which directly regulated BZR1-targeted genes expression in Col-0 and bri1-116.Comparing cold regulated genes,BZR1 binding genes and bzr1-D affected genes,14% bzr1-D affected genes were identified as BZR1-targeted genes,and more than half of these BZR1-targeted genes were regulated by cold.Combined with BZR1-targeted genes and cold-regulated genes of Col-0,bri1-116 and bin2-1,it was found that nearly 82% cold-regulated and BZR1-targeted genes were independent of BR signal,indicating that cold induction activates BZR1,and then regulates BZR1-targeted genes.3.Freezing test showed that BR biosynthesis or receptor-defective mutants get stronger constitutive freezing tolerance than Col-0,while gain-of-function bin2-1 are sensitivity to freezing stress.In addition,the constitutive freezing tolerance of bin2-1/bzr1-D was restored by bzr1-D,while bir1-116/bzr1-D are more sensitivity to freezing stress,indicating that the constitutive freezing tolerance of BR mutants was regulated by BZR1-dependent and BZR1-independent pathways.Under cold acclimation conditions,dwf4,bri1-116 and bin2-1 showed more sentitivity to freezing stress,while the freezing tolerance of bri1-116/bzr1-D and bin2-1/bzr1-D was restored by bzr1-D,indicating that the freezing tolerance of BR mutants was regulated by BZR1-dependent pathways.4.The relative length root of dwf4 and bri1-116 was promoted by low temperature.q PCR analysis showed that the expression of growth-associated BZR1-targeted genes was repressed by cold,but the repression of BZR1-targeted genes in Col-0 were severer than in bri1-116 and dwf4.Transcriptome analysis revealed that a large number of SAUR genes were induced by BR and repressed by cold.Overexpression of SAUR6,SAUR14 and SAUR16 promotes plant growth at room temperature.In addition,cold induction of SAUR6,SAUR14 and SAUR16 driven by the RD29 A promoter promoted the plant growth under low temperature conditions.Overexpression of SAUR16 and cold induction of SAUR16 driven by the RD29 A are sensitivity to freezing stress.These results suggest that BZR1 is activated by cold directly or indirectly inhibits the expression of growth-related genes,thereby coordinating plant growth and cold stress.Recent studies have shown that BR and its related transcription factor BZR1 are involved in cold responses of plant,and our findings confirmed these results.More importantly,the results of this study found that the role of BZR1 in cold responses of plant via a BR-independent manner,while the cold-activated BZR1 regulates the expression of some genes(such as SAURs)in a completely opposite way to BR.The results of this study can help reveal the complex roles of BR and its signaling pathway in cold responses of plant,providing new insights of freezing tolerance and growth mechanisms at low temperature.