葡萄HXK基因家族的鉴定与表达分析

Identification and Expression Analysis of Grape HXK Gene Family

为了对葡萄HXK基因家族进行鉴定和表达分析,利用拟南芥和玉米的HXK(Hexokinase)基因注册序列,从葡萄全基因组中鉴定得到HXK基因4个,并命名为VvHXK1、VvHXK2、VvHXK3和VvHXK4,对其进行生物信息学分析。蛋白质理化性质分析结果表明,每个基因编码的氨基酸个数分布在412-524 aa,VvHXK4是碱性蛋白,其余3个均为酸性蛋白;亚细胞定位分析表明,4个VvHXK基因都在细胞质中表达,且VvHXK4主要定位于叶绿体,而VvHXK2也存在于线粒体和细胞核中;蛋白质的二级结构预测表明,4个VvHXK基因编码的蛋白主要以α-螺旋为主;外显子结构分析表明,除VvHXK1含10个外显子外,其余3个都有9个外显子,说明VvHXK基因比较保守;启动子顺式作用元件分析表明,4个VvHXK基因均含有ABA响应元件和MYB及WRKY转录因子的结合区域,而VvHXK4中没有发现低温响应元件;系统进化树分析表明,VvHXK1与拟南芥AtHXK1的同源关系最近,推测其不仅具有调节生长发育,影响根系的生长等功能,而且具有加速衰老的作用。VvHXK2与马铃薯StHXK1的同源关系最近,推测其具有使叶绿体中的葡萄糖磷酸化活性升高的功能。VvHXK3与马铃薯StHXKRP1的同源关系最近,可能具有诱导离体叶片中糖表达的作用。VvHXK4与菠菜SoHXK1的同源关系最近。荧光定量分析表明,VvHXK2、VvHXK3和VvHXK4均在20 g/L的葡萄糖处理下相对表达量最高,即其对葡萄糖磷酸化最显著,在果糖和蔗糖的处理下表达量较低。

The purpose of this study was to identify and analyze HXK gene family in grape.Based on the HXK(hexokinase)gene registration sequence of Arabidopsis thaliana and maize,four HXK genes were identified from the whole genome of Vitis vinifera,namd as VvHXK1,VvHXK2,VvHXK3 and VvHXK4.The bioinformatics analysis were performed on these four genes.The results of protein physicochemical properties showed that the number of amino acids encoded by the four genes was between 412 aa and 524 aa,of which VvHXK4 was a basic amino acid,and the other three were acidic amino acids.In addition,subcellular localization analysis indicated that all four VvHXK genes were localized in the cytoplasm.VvHXK4 was mainly localized in chloroplasts,and VvHXK2 might appear in mitochondria and nucleus.Meanwhile,the secondary structure prediction of the proteins found that the proteins encoded by the four VvHXK genes were mainlyα-helix.The structural analysis of exons indicated that VvHXK1 contained 10 exons,and others contained 9 exons,showing that the VvHXK genes were relatively conserved;The initiation of cis-acting element analysis indicated that four VvHXK genes contained ABA response elements,MYB binding region and WRKY transcription factors,but no low temperature response component were found in VvHXK4.Phylogenetic tree analysis showed that the homology relationship between VvHXK1 and Arabidopsis AtHXK1 was the closest,indicating that it not only had the functions of regulating growth and development and affecting the growth of roots,but also had the effect of accelerating aging.The homology relationship between VvHXK2 and potato StHXK1 was recent,showing that it could increase the glucose phosphorylation activity in leaf extract or chloroplast.The homology relationship between VvHXK3 and potato StHXKRP1 was the closest,suggesting that VvHXK2 might be able to induce the expression of sugar in leaves.The homology relationship between VvHXK4 and spinach SoHXK1 was recent,which was important for studying the glucose metabolism of grapes.Fluorescence quantitative analysis showed that VvHXK2,VvHXK3 and VvHXK4 had the highest relative expression under the treatment of 20 g/L glucose,which was the most significant for glucose phosphorylation,and low expression under the treatment of fructose and sucrose.This is important for studying the sugar metabolism of grapes.