Identification Of Rice And Arabidopsis TFⅢA Zinc Finger Protein Families And Functional Analysis Of ZFP207

Rice is one of the most important crops worldwide,but rice growth and productivity are largely affected by abiotic stresses,including low temperature,drought and high salinity. A series of changes from cellular to physiological levels occur after rice plants are stressed. These genes in responses to abiotic stresses can be classified into two groups:one encoding the function proteins and osmoprotectants directly to protect plant cells against stresses, another encoding protein kinases and transcription factors,functioning in signal transduction and regulating gene expression.The zinc finger proteins represent a large transcription factor family in plant.It could be divided into several subfamilies including TFâ…¢A,WRKY,DOF,GATA,RING,and PHD,based on the various functions.Recently, many researches focused on the TFâ…¢A-type zinc finger genes with a conserved sequence "CX2CX3FX3QALGGHX3H" in zinc finger domains.About 40 TFâ…¢A-type zinc finger proteins have been isolate from plant,most of them play important roles in the developmental processes or responses to abiotic stresses in plant.In order to identify TFâ…¢A-type zinc finger genes from rice(Oryza sativa L.) and Arabidopsis,the amino acid sequence of several genes reported in this family,were used as query probes to search plant genome database through tBLASTn algorithm program.It was totally identified 72 and 59 of TFâ…¢A-type zinc finger protein genes in rice and Arabidopsis respectively.Analysis of the deduced amino acid sequences showed that most of these proteins encoded one or two zinc finger domains.These genes,in silicon,were randomly distributed on the 12 rice chromosomes and 5 Arabidopsis chromosomes respectively,with a few genes clustered.Phylogenetic analysis of these proteins and other reported proteins in plants showed that plant TFâ…¢A-type zinc finger proteins could he classified them as three subfamilies based on the numbers of zinc finger domains contained: Subfamilyâ… ,most of members with single zinc finger domain and the reported genes mostly involved in plant growth and development,Subfamilyâ…¡,most of members with two zinc finger domains and the reported genes mostly related to responses to abiotic stresses;and Subfamilyâ…¢,most of members with 3-5 zinc finger domains and the reported proteins involved in plant growth,development and other key metabolisms.Through a cDNA microarray technique the expression patterns of rice TFâ…¢A-type zinc finger genes were screened in rice seedlings under cold,drought and H₂O₂ treatments.We obtained the data of 45 genes.To compare with control the expressions of at least 27 genes were changed with more than 2-fold.Among them 14 were responded to H₂O₂,and 8 genes to all three treatments.One single zinc finger protein gene,named ZFP207,was isolated from the cDNA of rice seedlings treated by salt stress by silicon cloning and RT-PCR approach.ZFP207 encodes 188 amino acids with the calculated molecular mass of 20.72 KDa.It contains a Cys2/His2-type zinc finger domain with the plant specific "QALGGH" motifs.However,in the ZFP207 there was no other domains such as NLS,L-box,DLN-box/EAR-motif discovered in most plant TFâ…¢A-type zinc finger proteins.ZFP207,fused with the green fluorescent protein(GFP) reporter gene,was introduced into onion epidermis cells.The ZFP207-GFP fusion protein was targeted to the nucleus.By real-time RT-PCR the inducement of ZFP207 under treatments with salt,drought,cold stress and ABA were evaluated.We constructed the 35S:ZFP207 vector and transferred it into tobacco plants by in vivo infiltration,and then we obtained the ZFP207-overexpression transgenic plants.The 2# and 4# transgenic lines were used for stresses tolerances assay.The results indicated that ZFP207-overexpression transgenic lines increased significantly cold,drought and high salt stresses tolerance compared to the WT plants,but the germination of ZFP207-overexpressed seeds is insensitive to ABA.In addition,the proline contents in transgenic plants were obviously higher than that in WT plants under normal condition. And the expression of Osmotin enhanced in transgenic lines,while the expressions of 3 LEA genes showed no change in transgenic plants.Up to date,ZFP207 is the first reported TFâ…¢A-type single zinc finger gene involved in abiotic stresses in plants.More analysis of ZFP207 will help to widen our understanding of the relation between the structures and functions of TFâ…¢A-type zinc finger proteins in plants.