马铃薯绿原酸调控因子的筛选与功能鉴定  

Screening and functional identification of chlorogenic acid regulatory factors in potato

作　　者：周洪源 杨慧芹[1] 罗威 石振明 马玲 ZHOU Hong-Yuan;YANG Hui-Qin;LUO Wei;SHI Zhen-Ming;MA Ling(Key Laboratory of Yunnan Province for Biomass Energy and Environmental Biotechnology,Yunnan Normal University,Kunming 650000,Yunnan,China)

机构地区：[1]云南师范大学云南省生物质能与环境生物技术重点实验室,云南昆明650000

出　　处：《作物学报》2024年第7期1740-1749,共10页Acta Agronomica Sinica

基　　金：广东省基础与应用基础研究重大项目(2021B0301030004);云南省William John Lucas院士工作站(202105AF150028);国家自然科学基金项目“二倍体栽培种马铃薯RH中自交亲和基因的图位克隆”(32002032);云南省基础研究专项面上项目“薯肉龙葵素含量负调控因子StERF9基因的克隆和功能解析”(202201AT070037);昭通市千和农业科技开发有限公司“尚轶专家工作站”项目资助。

摘　　要：马铃薯是世界第三大粮食作物。绿原酸是马铃薯中最主要的酚类化合物,也是马铃薯抗虫抗病的物质基础之一,但过量绿原酸会影响薯块的口感。因此,培育地上部分高绿原酸含量而薯肉中低绿原酸含量的品种能很好的兼顾马铃薯抗病性及品质口感的需求。为了明确马铃薯中绿原酸的分子调控机制,本研究以绿原酸合成关键酶基因StHQT的启动子序列为诱饵,进行酵母单杂交文库筛选,鉴定出一个AT-hook基因家族的转录因子StAHL。进一步对StAHL的生物信息学特征、表达模式、蛋白质产物的亚细胞定位等信息进行了系统分析,并利用酵母单杂交和双荧光素酶报告试验等方法验证了StAHL与StHQT启动子之间的转录活性。结果表明StAHL基因的表达没有组织特异性,但根与花中的表达量相对较高,其蛋白产物包含AT-Hook、DUF296两个保守结构域,且定位于细胞核中。StAHL蛋白能直接作用于StHQT的启动子序列,并抑制其转录活性,表明StAHL可能通过抑制StHQT的表达来抑制马铃薯中绿原酸的积累。研究结果为全面揭示马铃薯绿原酸生物合成的分子机制奠定了基础,为马铃薯精准育种提供了分子靶标。Potato is the third largest food crop in the world.Chlorogenic acid is the most important phenolic compound in potato,and also one of the material basis of potato insect resistance and disease resistance.However,excessive chlorogenic acid can affect the taste of tuber,therefore,breeding varieties with high chlorogenic acid content in the ground part and low chlorogenic acid content in tuber flesh can well meet the needs of potato disease resistance and quality taste.In order to clarify the molecular regulation mechanism of chlorogenic acid in potato,we used the promoter sequence of StHQT,a key enzyme for chlorogenic acid synthesis,as bait to screen the yeast single hybridization library,and identified the transcription factor StAHL of an AT-hook gene family.Further,the bioinformatic characteristics,the relative expression patterns,and subcellular localization of protein products of StAHL were systematically analyzed,and the transcriptional activity between StAHL and StHQT promoters was verified by yeast single hybridization and double luciferase report experiments.The results showed that StAHL gene expression was not tissue specific,but relatively high in roots and flowers.StAHL protein products contained two conserved domains,AT-Hook and DUF296,and were localized in the nucleus.StAHL proteins acted directly on StHQT promoter sequences and inhibited transcriptional activity.This suggests that StAHL may inhibit the accumulation of chlorogenic acid in potato by inhibiting StHQT expression.The results laid a foundation for revealing the molecular mechanism of chlorogenic acid biosynthesis in potato,and provided a molecular target for the breeding practices in potato.

关 键 词：马铃薯 绿原酸 调控因子 STAHL AT-HOOK 

分 类 号：S532[农业科学—作物学]