高光胁迫下香果树叶片生理生化指标变化及相关基因差异表达和功能分析

Variations of physiological and biochemical indexes and differential expression and functional analyses on related genes in leaves of Emmenopterys henryi under high light stress

以光照强度54μmol·m^(-2)·s^(-1)为对照,对高光胁迫(光照强度180μmol·m^(-2)·s^(-1))下香果树(Emmenopterys henryi Oliv.)幼苗叶片的部分生理生化指标变化进行分析;采用RNA-seq技术筛选高光胁迫下的差异表达基因,并对其进行功能和表达分析及qRT-PCR验证。结果表明:高光胁迫下,香果树叶片的净光合速率、核酮糖1,5-二磷酸羧化酶/加氧酶活性和叶绿素含量显著下降,超氧化物歧化酶、过氧化氢酶、过氧化物酶和抗坏血酸过氧化物酶活性以及花青素和脱落酸含量显著升高。高光胁迫下,香果树的差异表达基因有6776个,其中上调和下调差异表达基因分别有3063和3713个。GO功能富集分析结果显示:上调和下调差异表达基因富集到的GO功能条目分别有2022和1835个,显著富集的GO功能条目分别有196和129个,基因功能均包括生物过程、分子功能和细胞组分3类。KEGG代谢通路富集分析结果显示:这些差异表达基因显著富集到次生代谢物生物合成、代谢途径、光合作用等20个代谢通路中。其中,光合作用相关基因Lhca4、Lhcb1、Lhcb2、Lhcb5、psaA、psaE、psaG、psaH、psaK、psaL、psaO、psbC、psbK、psbO、psbP、psbQ、psbR、psbW、petA、petC、petG、petE、atpC1、atpI、RBCS、rbcL、RCA、HEMA1、PORA以及脱落酸合成及信号通路相关基因NCED1、NCED3、CYP707A、PP2C51的表达水平显著下调;而光合作用相关基因psbS、atpA、atpD,花青素合成相关基因,抗氧化酶相关基因以及脱落酸合成及信号通路相关基因BGLU11、BGLU46、BGLU23、BGLU24、BGLU34、PYR/PYL、SnRK2C的表达水平显著上调。qRT-PCR验证结果显示:供试8个差异表达基因的表达水平与RNA-seq检测结果一致。综上所述,香果树主要通过调控光合作用、抗氧化酶系统、脱落酸代谢及信号通路以及花青素代谢通路响应高光胁迫。

Taking illumination intensity of 54μmol·m^(-2)·s^(-1)as the control,variations of some physiological and biochemical indexes in leaves of Emmenopterys henryi Oliv.seedlings under high light stress(illumination intensity of 180μmol·m^(-2)·s^(-1))were analyzed;differentially expressed genes under high light stress were screened by using RNA-seq technology,and their functional and expression analyses and qRT-PCR verification were conducted.The results show that under high light stress,net photosynthetic rate,ribulose 1,5-bisphosphate carboxylase/oxygenase activity,and chlorophyll content in leaves of E.henryi significantly decrease,activities of superoxide dismutase,catalase,peroxidase and ascorbate peroxidase,contents of anthocyanin and abscisic acid significantly increase.There are 6776 differentially expressed genes in E.henryi under high light stress,in which,up-regulated and down-regulated differentially expressed genes are 3063 and 3713,respectively.The GO functional enrichment analysis result shows that the up-regulated and down-regulated differentially expressed genes are enriched into 2022 and 1835 GO functional items,respectively,and there are 196 and 129 GO functional items enriched significantly,respectively,the gene function contains three categories namely biological process,molecular function,and cellular component.The KEGG metabolic pathway enrichment analysis result shows that these differentially expressed genes are significantly enriched into 20 metabolic pathways including biosynthesis of secondary metabolites,metabolic pathways,photosynthesis,etc.In which,the expression levels of photosynthesis-related genes Lhca4,Lhcb1,Lhcb2,Lhcb5,psaA,psaE,psaG,psaH,psaK,psaL,psaO,psbC,psbK,psbO,psbP,psbQ,psbR,psbW,petA,petC,petG,petE,atpC1,atpI,RBCS,rbcL,RCA,HEMA1,PORA and abscisic acid synthesis and signal pathway-related genes NCED1,NCED3,CYP707A,PP2C51 are significantly down-regulated,while those of photosynthesis-related genes psbS,atpA,atpD,anthocyanin synthesis-related genes,antioxidase-related genes,and abscisic acid synthesis and signal pathway-related genes BGLU11,BGLU46,BGLU23,BGLU24,BGLU34,PYR/PYL,SnRK2C are significantly up-regulated.The qRT-PCR verification result shows that the expression levels of eight test differentially expressed genes are consistent with the RNA-seq detection result.Taken together,E.henryi mainly responds to high light stress via regulating photosynthesis,antioxidase system,abscisic acid metabolism and signal pathway,and anthocyanin metabolic pathway.