The Molecular Mechanisms Of BpSZA1 From Betula Platyphylla In Modulating Drought And Salt Tolerance

Plants typically suffer multiple abiotic stresses including salt and drought during their life cycle.In abiotic stress condition,plants can regulate physiological and biochemical reactions by certain transcription factors to enhance abiotic stress tolerance.C2H2-type zinc finger protein（ZFP）play an important role in signaling pathways that regulate plant responses to abiotic stress.In this study,a C2H2 zinc finger protein gene was identified from the transcription data associated with salt stress in Betula platyphylla.Suk.,the gene was steadily up-regulated at all-time points under salt stress,and it was named BpSZA1（salt-induced zinc finger protein of Arabidosis thaliana,MZ544846）,and its function and regulation mechanism in responding to high salt and drought were preliminary studied.The main results are as follows:（1）A C2H2 zinc finger protein BpSZA1 was identified from B.platyphylla Suk.,BpSZA1protein is localized to the nucleus.The relative expression level of BpSZA1 in adult stem,old stem,old leaf,root,adult leaf and young leaf decreased successively.BpSZA1 was induced by abiotic stress including salt and drought and hormones including abscisic acid（ABA）,gibberellin（GA）and Methyl jasmonate（Me JA）.There are many stress response elements in BpSZA1 promoter region including GT1GMSCAM4,EBOXBNNAPA,MYBCORE and ABRELATERD1,and BpSZA1 promoter is a stress-induced promoter,meanwhile BpSZA1promoter responded to Na Cl,PEG and ABA treatment.（2）The overexpressing（OE）and RNAi-silencing（Ri）lines of BpSZA1 were generated by transgenic technique.Under drought and salt condition,the growth parameters including plant height,fresh height and root length and growth status of OE lines plants were significantly better than those of WT,however those of Ri plants were significantly worse than those of WT.In addition,overexpression BpSZA1 decreased MDA and H₂O₂contents and Electrolyte leakage,increased SOD,POD activities and chlorophyll and proline contents.ABA induced stomatal closure in OE lines.These results indicated that BpSZA1 positively regulates drought and salt tolerance of B.platyphylla.（3）RNA-seq analysis was perform by using OE6,Ri7 and WT birch,there are 884up-regulated genes and 716 down-regulated genes were identified under control condition;and there are 1119 up-regulated genes and 1841 down-regulated genes were identified under salt-treated condition.Among differentially expressed genes（DEGs）,including many genes including involved in abiotic stress response.（4）By software prediction,yeast one-hybrid experiment and co-transformation experiment of effector vector and reporter vector experiment verify that BpSZA1 protein could bind to the element which core sequence is GAGAGA.The result of Ch IP-PCR and Ch IP-q PCR furher demonstrated that,BpSZA1 protein bound to the promoter regions of Bp APX1,Bp APX2,Bp CAT and Bp6PGDH genes,and regulated the expression of these genes.（5）Transcriptional activation experiments showed that BpSZA1 gene has transcriptional activation activity,and its transcriptional activation region is located at C-terminal 212-247amino acids.The protein Bp DH interacting with BpSZA1 was identified by Y2H assay between BpSZA1 and B.platyphylla c DNA library.Through bimolecular fluorescence complementation（Bi FC）validation,BpSZA1 could interact with Bp DH.Overexpression Bp DH enhanced the salt tolerance of plants by increasing reactive oxygen species（ROS）scavenging and proline content.In conclusion,this study preliminarily explored the mechanism of BpSZA1 in drought and salt tolerance.On the one hand BpSZA1 positively regulates drought and salt tolerance by binding to GAGAGA elements in the promoter region of downstream target genes including Bp APX1,Bp APX2,Bp CAT and Bp6PGDH.On the other hand,BpSZA1 protein could interact with Bp DH to regulate salt and drought tolerance.In this study,we identify a gene induced by drought and high salinity,and preliminarily explored its regulation mechanism of high salinity and drought stress,providing a theoretical basis for understanding the mechanism of drought and salt tolerance and breeding of B.platyphylla...