Preliminary Analysis Of Salt Tolerance Function Of BpHsfB1 Transcription Factor From Betula Platyphylla

Heat shock transcription factor(Hsf)has been proved to be an important transcriptional regulatory gene in plant cells,which plays an important role in plant resistance to heat stress and other life activities.In this study,a BpHsfB1 gene was cloned from Betula Platyphylla and its salt tolerance function was studied.The BpHsfB1 gene was 975 bp in length.The results of mulit-sequences alignment showed that the protein had high homology with the Hsf family of Arabidopsis.The phylogenetic tree showed that it belongs to the same family as the AT4G6990 of Arabidopsis,and belongs to Class B of Hsf family.So this gene was named BpHsfB1.The 1559bp promoter of BpHsfB1 was obtained from the genomic sequence of birch.The promoter sequence contained ABRE,TGA-element and ACE cis-regulatory element,which were related to plant stress resistance,auxin response and light response.The results of subcellular localization showed that BpHsfB1 protein was expressed in the nucleus.The analysis of transcriptional activation activity showed that BpHsfB1 transcription factor had no transcriptional activation activity,which might play a role through interaction with other transcription factors.The expression levels of BpHsfB1 in the roots,stems and leaves of birch under seven different stress conditions showed that the expression of BpHsfB1 was changed at different degrees after stress in the roots,stems and leaves of birch,which suggested that BpHsfB1 might be involved in many abiotic stress responses.Especially after salt stress,the expression of BpHsfB1 changed obviously.After 24 h,the expression level in stem and leaf began to increase,and the expression level in root,stem and leaf increased significantly at 48 h,the highest expression level was 5.07 times of the control.BpHsfB1 may play an important role in the salt tolerance of Birch.In order to further analyze whether BpHsfB1 gene was involved in salt stress response,the overexpression vector of BpHsfB1(pROKII-BpHsfB1)was constructed,and the overexpression strain of BpHsfB1 was obtained by agrobacterium-mediated method.Interestingly,multiple collateral branches were found in over-expressing the BpHsfB1 birch.Morevore,after 17 days of 120 mM NaCl stress,the control birch basically withered,but the over-expressed BpHsfB1 lines showed that some leaves withered but new branches and leaves grew.In addition,the activities of POD,SOD,MDA and H2O2 in transgenic and control birch under salt stress were measured and compared.The results showed that the activities of POD and SOD of the over-expressed BpHsfB1 birch were significantly higher than those of the control.And the contents of H2O2 and MDA were significantly lower than those of the control.These results indicated that the over-expression of BpHsfB1 gene had little damage after salt stress,and over-expression of BpHsfB1 gene could improve the salt tolerance of transgenic birch.Hsf transcription factors regulate the downstream gene expression by identifying Heat Shock Element(HSE)and participate in plant life activities.In this study,we screened for BpHsfB1 gene recognition in birch by Yeast one-hybrid and identified the selected components using Plant Care.The results showed that,except for HSE(AAAATTTC)elements,the BpHsfB1 protein may also recognize the ABRE(ACCGTGACGTG)element.Further the results of yeast one-hybrid and EMSA demonstrated that the BpHsfB1 protein of birch can specifically recognize both HSE element and ABRE.The results of ChIP-PCR showed that the BpHsfB1 transcription factor could bind ABRE elements on the promoters of SOD,POD,P5CS and SOS,and enhance the expression of the downstream genes in Birch.Whether BpHsfB1 transcription factor can regulate the expression of more downstream genes remains to be proved in the future.