摘要
包含SPX、SPX-EXS、SPX-MFS和SPX-RING四个亚族的植物 SPX基因家族在磷信号应答中发挥重要功能,但迄今对小麦的该家族基因成员功能了解尚少.本研究前期从小麦(Triticum aestivum)中鉴定得到一个SPX亚族成员基因TaSPX1(GenBank No.Ak332300),亚细胞定位分析发现其定位于细胞核.对TaSPX1 和来自小麦、拟南芥和水稻 SPX 家族的同源蛋白进行系统进化分析,结果表明,其与水稻 SPX 亚族的 OsSPX1 亲缘关系较近.应用RT-qPCR 技术研究发现,TaSPX1 的表达量在低氮胁迫下显著增加.构建烟草(Nicotiana tabacum)过表达转基因系(overexpression lines,OE),应用MS营养液培养对野生型(WT)和OE株系OE3 和OE4 植株表型进行鉴定.发现在低氮胁迫下,OE3 和 OE4 较 WT表现明显的生长优势,植株鲜重、根重和叶面积显著增加;包括光合速率、胞间 CO2浓度、气孔导度和蒸腾速率在内的光合参数,以及氮含量、可溶性糖、可溶性蛋白和叶绿素含量也都较 WT 显著增加.对氮吸收和同化相关基因的表达和酶活性测定结果表明,上述基因的部分成员在OE植株中的表达量和氮同化酶活性升高.此外,对包括SOD、POD和CAT在内的植株活性氧清除相关酶活力和MDA含量测定表明,与WT相比,OE植株中保护酶的活性均明显提高,MDA含量降低.同时发现OE植株中部分保护酶基因成员的表达水平也较WT明显升高.这些结果初步证实了 TaSPX1 通过改善光合参数、增强氮吸收和转运以及加强保护酶系统等在介导植株抵御低氮胁迫中发挥重要作用.本项研究对小麦SPX家族成员抵御非生物逆境功能增加了新认识,为作物抗低氮营养逆境的遗传改良提供了理论依据.
Abstract
The SPX gene family includes four subgroups:SPX,SPX-EXS,SPX-MFS,and SPX-RING,which play an important role in phosphate signal response,but so far,little is known about the functions of this family in wheat.Previously,we identified a gene TaSPX1(GenBank No.Ak332300),belonged to SPX subfamily from wheat(Triticum aestivum).Subcellular localization analysis showed that it targeted onto nucleus.Phylogenetic tree of TaSPX1 and its homologous proteins from the wheat,Arabi-dopsis,and rice SPX families showed that it was closely related to OsSPX1,a member of rice SPX subfamily.The relative ex-pression level of TaSPX1 significantly increased under low nitrogen(low-N)stress when investigated by RT-qPCR.Transgenic tobacco(Nicotiana tabacum)overexpression lines were generated.Using the culture methods of Murashige&Skoog(MS)hydroponic solution,the phenotype of WT and OE under low-N stress treatment was investigated.We found that the plants of OE3 and OE4,two OE lines overexpressing TaSPX1,displayed increased growth vigor and leaf area,together with the enhanced plant fresh weight and root weight,and elevated photosynthetic parameters including photosynthetic rate(Pn),intercellular carbon dioxide concentration(Ci),stomatal conductance(Gs),and transpiration rate(Tr),along with the increased contents of nitrogen,soluble sugar,soluble protein,and chlorophyll content upon low-N stress with respect to WT.Studies on transport and assimila-tion related parameters showed that under low-N stress,the relative expression level of some related genes and the activities of nitrogen assimilation-related enzymes were increased.Assays on the SOD,POD,and CAT,the enzymes functional as cellular protector,revealed the higher activities of them in OE plants than those in WT.On the contrary,MDA content was decreased.Further RT-qPCR analysis indicated the expression levels of several protection enzymes mentioned above were higher in OE plants than those of WT under low-N stress.Therefore,TaSPX1 played an important role in mediating plant resistance to low-N stress by improving photosynthetic parameters,enhancing nitrogen absorption and transport,and strengthening the protective enzyme system.The results enrich new understanding on the function of wheat SPX family members involved in abiotic stress,and provide a theoretical basis for genetic improvement of crops against low-N stress.
维普
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