外源氨基酸对拟南芥EBP1蛋白质合成影响的初步分析

Preliminary Analysis of the Effect of Exogenous Amino Acids on the Synthesis of EBP1 Protein in Arabidopsis thaliana

【目的】探讨植物吸收外源氨基酸的种类以及这些氨基酸对植物体内EBP1蛋白质合成的影响。【方法】本研究建立如下体系:在缺氮培养基中,分别添加16种终浓度为3 mmol/L的氨基酸来处理拟南芥Col-0。用可以恢复Col-0缺氮表型的氨基酸处理35S::EBP1-GFP报告植物后,观察荧光信号并结合核糖体提取及蛋白质印迹法(Western-blot)检测EBP1在mRNA及蛋白水平的表达。【结果】精氨酸、丙氨酸、天冬酰胺、谷氨酰胺、天冬氨酸和甘氨酸等6种氨基酸可以恢复Col-0的缺氮表型,同时也表明建立的体系切实可行。荧光观察试验表明:甘氨酸、丙氨酸、精氨酸和谷氨酰胺能促进35S::EBP1-GFP的荧光积累;核糖体提取试验表明:精氨酸、丙氨酸和甘氨酸能在翻译层面促进EBP1的蛋白质积累。Western-blot试验表明:丙氨酸和谷氨酰胺在蛋白水平上促进EBP1蛋白质的积累。【结论】精氨酸、丙氨酸、天冬酰胺、谷氨酰胺、天冬氨酸和甘氨酸可作为潜在的有机氮源供植物吸收利用,丙氨酸能同时在翻译和蛋白质层面促进EBP1的蛋白质积累。氨基酸被植物吸收后通过影响植物体内蛋白质的合成影响植物的生长发育。

[Purpose]To investigate the types of exogenous amino acids absorbed by plants and the effect of these amino acids on the synthesis of EBP1 protein in plants.[Method]We established the following system,Col-0 were grown on the 1/2 nitrogen deficient medium amended with 16 kinds of amino acids(3 mmol/L),respectively.Then,we treated the report plant 35S::EBP1-GFP with the amino acids which could be used as potential organic nitrogen sources to observe their fluorescence signals and combined with ribosome extraction essay and Western-blot essay to detect the expression of EBP1 in mRNA level and protein level,respectively.[Result]There were the kinds of amino acids including arginine,alanine,asparagine,glutamine,asparagine and asparagine which could be used as potential organic nitrogen sources.It also showed that the established system was feasible.The fluorescence observation essay showed that glycine,alanine,arginine and glutarnine could promote the fluorescence signals of 35S::EBP1-GFP;the ribosome extraction essay showed that glycine,alanine and arginine significantly promoted the mRNA expression of EBP1;and the Western-blot essay showed that alanine and glutarnine promoted the accumulation of EBP1 protein at the protein level.[Conclusion]Arginine,alanine,asparagine,glutamine,aspartic acid and glycine can be used as potential organic nitrogen sources for plant absorption and utilization.Moreover,alanine can promote the protein accumulation of EBP1 at the mRNA and protein levels.Exogenous amino acids regulate the growth and development of plants by affecting proteins synthesis.