甘薯蔗糖转运蛋白的功能分析  

Functional analysis on sucrose transporters in sweet potato

作　　者：刘怡然 吴正丹[1,2,4] 吴维泰 杨朝彬 陈才睿 张凯 LIU Yiran;WU Zhengdan;WU Weitai;YANG Chaobin;CHEN Cairui;ZHANG Kai(College of Agronomy and Biotechnology,Southwest University,Chongqing 400715,China;Chongqing Key Laboratory of Biology and Genetic Breeding for Tuber and Root Crops,Chongqing 400715,China;Engineering Research Center of South Upland Agriculture,Ministry of Education,Chongqing 400715,China;Cash Crops Research Institute,Guangxi Academy of Agricultural Sciences,Nanning 530007,Guangxi,China)

机构地区：[1]西南大学农学与生物科技学院,重庆400715 [2]薯类生物学与遗传育种重庆市重点实验室,重庆400715 [3]南方山地农业教育部工程研究中心,重庆400715 [4]广西壮族自治区农业科学院经济作物研究所,广西南宁530007

出　　处：《生物工程学报》2023年第7期2772-2793,共22页Chinese Journal of Biotechnology

基　　金：国家重点研发计划(2018YFD1000705,2018YFD1000700);中央高校基本科研业务费专项资金(XDJK2020B032,XDJK2021F001);重庆市技术创新与应用发展专项重点项目(cstc2021jscx-gksbX0022);西南大学种质创制专项研究项目。

摘　　要：甘薯(Ipomoea batatas)是重要的粮食和工业加工原料作物。蔗糖是植物体内碳水化合物长距离转运的主要形式,蔗糖转运蛋白(sucrose transporter,SUT)在植物的生长代谢中调控蔗糖的跨膜运输和分配,在韧皮部介导的源-库蔗糖运输和为库组织供应蔗糖的生理活动中起关键作用。本研究根据不同淀粉性状甘薯块根中差异表达的2个SUT基因转录本,进行cDNA末端快速扩增(rapid amplification of cDNA ends,RACE)克隆,获得IbSUT62788和IbSUT81616的全长cDNA序列;通过系统发育分析明确其分类;通过在本氏烟草(Nicotiana benthamiana)中瞬时表达明确其亚细胞定位;通过酵母功能互补系统鉴定IbSUT62788和IbSUT81616是否具有吸收、转运蔗糖和己糖的能力。通过实时荧光定量PCR(real-time fluorescence quantitative polymerase chain reaction,RT-qPCR)分析IbSU62788和IbSUT81616在甘薯各器官中的表达特征;通过蘸花法得到外源表达IbSUT62788和IbSUT81616基因的拟南芥(Arabidopsis thaliana)植株,比较与野生型拟南芥的淀粉和糖含量的差异。结果表明,IbSUT62788和IbSUT81616分别编码505个和521个氨基酸的SUT蛋白,均属于SUT1亚家族。IbSUT62788和IbSUT81616均定位于细胞膜,在酵母系统中具有转运蔗糖、葡萄糖和果糖的能力。此外,IbSUT62788还具有转运甘露糖的能力。IbSUT62788在甘薯叶片、侧枝和茎中的表达量更高,IbSUT81616在侧枝、茎和块根中表达量更高。IbSUT62788和IbSUT81616在拟南芥中异源表达后,植株可以正常生长,但生物量增加。IbSUT62788的异源表达增加了拟南芥植株叶片可溶性糖含量、叶片大小和种子千粒重;IbSUT81616的异源表达增加了拟南芥植株叶片、根尖的淀粉积累量和种子千粒重,但减少了可溶性糖含量。本研究结果表明,IbSUT62788和IbSUT81616可能是调控甘薯蔗糖和糖含量性状的重要基因,在细胞膜上进行着蔗糖的跨膜运输、蔗糖进出库组织、韧皮部蔗糖Sweet potato is an important food crop that can also be used as an industrial raw material.Sucrose is the main form of long-distance carbohydrate transport in plants,and sucrose transporter(SUT)regulates the transmembrane transport and distribution of sucrose during plant growth and metabolism.Moreover,SUT plays a key role in phloem mediated source-to-sink sucrose transport and physiological activities,supplying sucrose for the sink tissues.In this study,the full-length cDNA sequences of IbSUT62788 and IbSUT81616 were obtained by rapid amplification of cDNA ends(RACE)cloning according to the transcripts of the two SUT coding genes which were differentially expressed in sweet potato storage roots with different starch properties.Phylogenetic analysis was performed to clarify the classification of IbSUT62788 and IbSUT81616.The subcellular localization of IbSUT62788 and IbSUT81616 was determined by transient expression in Nicotiana benthamiana.The function of IbSUT62788 and IbSUT81616 in sucrose and hexose absorption and transport was identified using yeast functional complementarity system.The expression pattern of IbSUT62788 and IbSUT81616 in sweet potato organs were analyzed by real-time fluorescence quantitative PCR(RT-qPCR).Arabidopsis plants heterologous expressing IbSUT62788 and IbSUT81616 genes were obtained using floral dip method.The differences in starch and sugar contents between transgenic and wild-type Arabidopsis were compared.The results showed IbSUT62788 and IbSUT81616 encoded SUT proteins with a length of 505 and 521 amino acids,respectively,and both proteins belonged to the SUT1 subfamily.IbSUT62788 and IbSUT81616 were located in the cell membrane and were able to transport sucrose,glucose and fructose in the yeast system.In addition,IbSUT62788 was also able to transport mannose.The expression of IbSUT62788 was higher in leaves,lateral branches and main stems,and the expression of IbSUT81616 was higher in lateral branches,stems and storage roots.After IbSUT62788 and IbSUT81616 were heterologously

关 键 词：甘薯 蔗糖转运蛋白 可溶性糖 淀粉 

分 类 号：S566.1[农业科学—作物学] Q943.2[生物学—植物学]