Study On The Mechanism Of C2H2 Zinc Finger Protein TaZFP21 Regulating Drought Stress In Triticum Aestivum L.

Bread wheat(Triticum aestivum L.)is one of the major grain crops.The growth,development and yield of wheat are adversely affected by multiple abiotic stresses,such as drought,salt,high temperature and low temperature.In the process of plant adaptation to stress,plants recognize the changes of environmental factors through specific intracellular receptors.It can regulate the expression of related genes and plant growth and development by corresponding signal transduction pathways.The corresponding physiological and biochemical changes are produced in the plant,so that plants gradually adapt to the growth under the adverse environment.In this study,we aim to elucidate the molecular mechanism of TaZFP21 regulating the response to drought stress in wheat.Firstly,the comprehensive analyses the characteristics of wheat C2H2-ZF genes family.Secondly,The overexpression lines of TaZFP21 in Arabidopsis and wheat were constructed,and their tolerance was analyzed under abiotic stresses,respectively.The molecular mechanism of TaZFP21 regulating wheat drought response was deeply analyzed,and the following main research results were obtained:(1)The 204 TaC2H2-type zinc finger proteins(TaC2H2-ZFP)family members were systematically analyzed.TaC2H2-ZFPs were divided into 10 different subgroups.Synteny analyses showed that TaC2H2-ZF gene was closely related to C2H2-ZF gene in Poaceae.The specific conserved motifs of the TaC2H2-type zinc finger domains were classified,NPL-and SFP1-box were firstly found to be located in C2H2-ZFP proteins.(2)qRT-PCR showed that both TaZFP21 responded to drought stress,which was consistent with the report that TaZFP21 was significantly up-regulated under drought stress.TaZFP21 was found to have two highly conserved QALGGH domains and an EAR motif by the analysis of amino acid sequence.It was verified that TaZFP21 had no transcriptional activation activity by yeast transcriptional self-activation and GUS activity experiments.The TaZFP21 was verified that was a zinc finger protein transcriptional repressor.(3)Wild-type Col-0 and Arabidopsis overexpression lines were treated with different concentrations of mannitol,NaCl and ABA.It was found that the germination rate,green bud rate and seedling tolerance of overexpression lines were significantly lower than those of wild type(WT)by the statistics.Moreover,it was found that the expressions of some stress-related genes(SOS2,DREB2C,RD22,RD29A,RD29B,POD,CAT,APX and AtP5CS)were significantly inhibited in TaZFP21 overexpressing plants.The present study showed that compared with wild type,TaZFP21 heterologous overexpression increased the sensitivity of Arabidopsis to drought,salt and exogenous ABA.(4)Wild-type wheat Fielder and TaZFP21 overexpressing wheat were treated by drought.The activities of ROS-related scavenging enzymes in TaZFP21 overexpressing plants were lower than the wild-type Fielder plants.This indicated that drought stress caused disturbance of ROS homeostasis in TaZFP21 overexpressing wheat.The transgenic plants of overexpressing of TaZFP21 gene reduced yield in wheat.The results showed that TaZFP21 overexpression resulted in the decreased wheat drought tolerance and yield.(5)Three(motif 13,motif 16,and motif 22)binding sites of TaZFP21 were predicted by ChIP-seq experiment.The EMSA experiment further confirmed that TaZFP21 can specifically bind to 3 motifs.According to the combined analysis of RNA-Seq and ChIP-seq,the expression of Tarbo-1p,TaAPX,TaCAT,TaSnRK2.5,TaSnRKγ1,and TaPro were significantly inhibited.The results showed that TaZFP21 may reduces the drought tolerance of wheat by inhibiting the expression of drought related to genes.TaZFP21 can specifically bind to the promoters of 6 target genes were verified by Y1H.Transcriptional inhibition activity was verified by LUC verify.The results showed that TaZFP21 may negatively regulating drought tolerance of wheat by inhibited the expression of 6 target genes.In conclusion,this study systematically analyzed the characteristics of wheat C2H2 zinc finger protein gene family and TaZFP21 zinc finger protein.It revealed a new mechanism of TaZFP21 negatively regulating wheat drought tolerance by inhibiting downstream gene expression.This study is great significance to further analyze the molecular mechanism of wheat drought resistance,and provides new genetic resources for wheat drought resistance molecular design and breeding.