雨生红球藻蓝光受体向光素(PHOT)互作蛋白鉴定及烟草异源表达的生理效应  

ldentification of blue light receptor phototropin(PH0T)interacting protein from Haematococcus pluvialis and physiological effect of its heterologous expression in tobacco

作　　者：李旺宁 李亚男 梁梦静 张春辉 季春丽 李润植 崔玉琳[2] 秦松[2] 薛金爱 崔红利 LI Wangning;LI Yanan;LIANG Mengjing;ZHANG Chunhui;JI Chunli;LI Runzhi;CUI Yulin;QIN Song;XUE Jin'ai;cUI Hongli(College of Agronomy/lnstitute of Molecular Agriculture and Bioenergy,Shanxi Agricultural University,Jinzhong,Shanxi 030801,China;Key Laboratory of Coastal Biology and Biological Resources Utilization,Yantai Institute of Coastal Zone Research,Chinese Academy of Sciences,Yantai,Shandong 264003,China)

机构地区：[1]山西农业大学农学院/分子农业与生物能源研究所,山西晋中030801 [2]中国科学院烟台海岸带研究所海岸带生物学与生物资源保护重点实验室,山东烟台264003

出　　处：《植物生理学报》2023年第7期1300-1310,共11页Plant Physiology Journal

基　　金：国家自然科学基金(31902394和41876188);山西省高等学校科技创新计划(2021L119);山西农业大学科技创新基金(2018YJ16);优秀博士毕业生来晋工作奖励资金(SXYBKY2019036)。

摘　　要：向光素(PHOT)是植物和绿藻特有的蓝光受体,鉴定PHOT的互作蛋白及探究其功能为阐明PHOT介导的信号途径与生理功能奠定基础。本研究采用酵母双杂交技术从雨生红球藻cDNA文库中筛选HaePHOT的候选互作蛋白,共获得23个阳性克隆,其中4个克隆注释为DnaJ。利用RNA-Seq数据,对雨生红球藻中HaePHOT与DnaJ的表达模式进行相关性分析,发现二者呈很强的正相关;进一步分析HaePHOT或DnaJ与虾青素合成及积累的关键基因(PSY、PDS、LYCB、BKT、BCH、CYP97B、KAS、FAD)表达量的相关性,发现HaePHOT及DnaJ分别与LYCB、BKT、BCH、CYP97B呈正相关,而与PSY、PDS、KAS、FAD呈负相关。瞬时转化烟草结果显示,在正常光(白光)下,野生烟草和过表达HaePHOT的烟草植株叶绿素a、叶绿素b和类胡萝卜素差异不显著;但蓝光下差异显著,且转基因烟草色素含量较高。qRT-PCR结果显示叶绿素a加氧酶(CAO)、原叶绿素酸酯氧化还原酶(POR)、八氢番茄红素合酶(PSY)、八氢番茄红素脱氢酶(PDS)和番茄红素β-环化酶(LYCB)编码基因在蓝光照射及转基因烟草中的表达显著提高,均与叶绿素和类胡萝卜素含量的变化趋势一致。这些发现为后续解析HaePHOT及其互作蛋白介导的调控机制提供了理论基础。Photropin(PHOT)is a unique blue light receptor in plants and green algae.Identification of PHOT interacting proteins and their functions will lay a foundation for elucidating PHOT-mediated signaling pathways and physiological functions.In this study,the yeast two-hybridization(Y2H)was used to screen the candidate interacting proteins of HaePHOT from the cDNA library of Haematococcus pluvialis.A total of 23 positive clones were obtained,of which four clones were annotated as DnaJ.The expression correlation of HaePHOT and DnaJ in H.pluvialis showed a strong positive model basing on RNA-Seq data.Further analysis of the expression correlation between HaePHOT or DnaJ and astaxanthin synthetic genes showed that HaePHOT and DnaJ were positively correlated with LYCB,BKT,BCH and CYP97B,but negatively correlated with PSY,PDS,KAS and FAD,respectively.Transient expression assay in tobacco showed that there was no significant difference in chlorophyll a,chlorophyll b and carotenoids contents between wildtype tobacco and HaePHOT-overexpressing tobacco under normal light(white light).However,under blue light,the contents of those pigments were higher in the transient-expression plants than that in the wildtype tobacco.qRT-PCR analysis showed that the expression of genes encoding chlorophyll a oxygenase(CAO),protochlorophyllide oxidoreductase(POR),phytoene synthase(PSY),phytoene desaturase(PDS)and lycopeneβ-cyclase(LYCB)were significantly increased under blue light irradiation in the transient-expression tobacco plants,which was consistent with the change pattern of chlorophylls and carotenoids contents.These findings provide a theoretical basis for further revealing the regulatory mechanisms mediated by HaePHOT and its interacting proteins.

关 键 词：雨生红球藻(Haematococcus pluvialis) 向光素(PHOT) 互作蛋白 烟草异源表达 生理效应 

分 类 号：Q943.2[生物学—植物学]