Functional Analysis Of UV-B Photoreceptor BpUVR8 In Betula Platyphylla.

As a photoreceptor specifically for UV-B light,UVR8 gene plays an important role in the photomorphogenesis and developmental growth of plants.In this research,we isolated the UVR8 gene from birch,named BpUVR8(AHY02156),its expression patterns in different tissues and under different abiotic stress and hormone treatments were investigated.Meanwhile,the promoter sequence of BpUVR8 was obtained.The bioinformatics analysis and promoteractivity analysis were carried out.In addition,we studied the subcellular localization of the BpUVR8.We obtained the transgenic Arabidopsis of the Betula platyphylla BpUVR8 in the background of the wild type and the Arabidopsis uvr8 mutant,and identified the function of the BpUVR8 gene using functional complementation assay.In addition,the role of BpUVR8 gene in ABA signaling pathway was studied.The main results are as follows:1.Bioinformatics Analysis of BpUVR8 GeneThe bioinformatics analysis showed that BpUVR8 sequence consists of 1326bp with a complete open reading frame and encode 441 amino acids.Furthermore,it is a stable hydrophobic protein.This protein may not exist signal peptide but exist the transmembrane ability.The protein has two transmembrane domains.According to our tertiary structure prediction of the protein,it is constituted by a seven-bladed ?-propeller,which has a high structural homology with the AtUVR8 in Arabidopsis thaliana.2.Expression pattern in birchThe results of qPCR showed that BpUVR8 gene was mainly expressed in leaves,which was about 6.5 times of inflorescence.In addition,the expression of BpUVR8 gene was the highest in August,about twice as high as in June,which suggested that the expression of BpUVR8 gene was tissue-specific.Besides,the abiotic stress and ABA hormone can induce the expression of BpUVR8 gene.The results of subcellular localization showed that BpUVR8 protein was located in the nucleus and cell membrane.3.Bioinformatics and activity Analysis of the Promoter of BpUVR8 GeneWe obtained the promoter fragment of 2397 bp by SiteFinding-PCR method.The bioinformatics analysis of the promoter sequence showed that the promoter contained multiple light response elements,but also a number of tissue-specific cis elements involved in the hormone stress and biotic and abiotic stress.The promoter of BpUVR8 gene was linked to the reporter gene GUS,and the recombinant vector pBpUVR8::GUS was transferred into tobacco instantly.The promoter of BpUVR8 gene was found to have promoter activity by GUS staining on transgenic tobacco.The expression of GUS gene in transgenic seedlings was induced under UV-B?NaCl?ABA and high temperature stress treatment,which indicated that BpUVR8 promoter not only has the promoting activity but can also respond to the induction of abiotic stress and ABA signal.4.Functional complementation assay of BpUVR8 in Arabidopsis uvr8 mutantsThe functional complementation test showed that BpUVR8 overexpression rescued the uvr8 mutant phenotype.This indicates that BpUVR8 is a positive regulator of UV-B-induced photomorphogenesis in plants.5.The role of BpUVR8 gene in ABA signaling pathwayBpUVR8 overexpression in plants indicated hypersensitivity to ABA under both white light and UV-B radiation at the stage of seed germination and growth.Moreover,BpUVR8 regulates the expression of a subset of ABA-responsive genes,both in Arabidopsis and Betula platyphylla,under the ABA treatment.