巴西橡胶树大、小橡胶粒子在乙烯调控天然橡胶合成中的作用分析

Role of Large and Small Rubber Particles of Hevea Brasiliensis in Ethylene Regulated Natural Rubber Synthesis

巴西橡胶树(Heveabrasiliensis)是重要的产胶植物,胶乳中的橡胶粒子(RP)是合成天然橡胶的重要细胞器,但其响应外源乙烯刺激调控天然橡胶合成的具体机制还不清楚。为了明确不同大小RP在此过程中的作用,本研究通过分离外源乙烯刺激后胶乳中不同直径的RP,并与对照组进行差异蛋白质组学分析。结果发现:在直径较大的大橡胶粒子(LRP)中鉴定出响应乙烯刺激差异蛋白37个,分别参与天然橡胶合成、糖酵解/糖异生、碳代谢及氨基酸生物合成等代谢通路,其中包含4个REF/SRPP家族成员;在直径较小的小橡胶粒子(SRP)中鉴定出56个差异蛋白,分别参与内质网中的蛋白质加工、内吞及剪接体等代谢通路,其中包含5个REF/SRPP家族成员。关键差异蛋白REF138具有较多等电点(pI)和分子量不同的蛋白亚型,LRP中低于标准等电点(4.80)的REF138亚型响应乙烯刺激积累减少,高于标准等电点的REF138亚型响应乙烯刺激积累增加;与LRP不同,SRP上有更多的REF138亚型响应乙烯刺激发生变化,具有标准分子量(14.7kDa)的REF138亚型响应乙烯刺激积累增加,高于标准分子量的亚型响应乙烯刺激积累减少。针对关键差异蛋白的互作蛋白功能分析发现,REF/SRPP家族成员REF138、REF175、REF258、SRPP117及SRPP204间存在相互作用,可能形成蛋白复合体结合在RP上。除REF/SRPP家族成员外,REF138的互作蛋白主要参与剪接体及内吞代谢通路,REF258的互作蛋白主要参与脂代谢过程与次生代谢产物合成调控。综上,通过对LRP及SRP上响应乙烯刺激差异蛋白及其互作蛋白的功能分析初步揭示了LRP和SRP响应外源乙烯刺激调控天然橡胶合成的代谢调控机制。

Hevea brasiliensis is an important natural rubber-producing plant.Rubber particles(RP)in latex are important organelles for the synthesis of natural rubber,which can be divided into large rubber particles(LRP)and small rubber particles(SRP)according to the diameter.Although the specific mechanism of its regulation of natural rubber synthesis in response to exogenous ethylene stimulation has been studied.However,the specific regulatory mechanism of LRP and SRP in regulating natural rubber synthesis in response to ethylene stimulation is still unclear.In order to clarify the role of RP with different diameters in the synthesis of natural rubber,LRP and SRP stimulated by ethylene were isolated and the proteins of corresponding samples were extracted for differential protein analysis.37 differential proteins in response to ethylene stimulation were identified in the LRP,which were involved in natural rubber biosynthesis,glycolysis/gluconeogenesis,carbon metabolism and amino acid biosynthesis,and some other metabolic pathways,including four members of the REF/SRPP family.56 differential proteins were identified in SRP,which were involved in protein processing,endocytosis,splicing in endoplasmic reticulum,and some other metabolic pathways,including five members of the REF/SRPP family.The key differential accumulation protein REF138 had many isoforms with different isoelectric points(pI)and molecular weights(MW).REF138 isoforms below the standard pI(4.80)on LRP were down-regulated in response to ethylene stimulation,while isoforms above the standard pI were up-regulated to ethylene stimulation.Different from LRP,more isoforms of REF138 on SRP changed in response to ethylene stimulation,and isoforms with standard MW(14.7 kDa)were up-regulated in response to ethylene stimulation,while isoforms with higher than standard MW were down-regulated in response to ethylene stimulation.The function analysis of key differential proteins showed that there were interactions among the members of the REF/SRPP family.REF138,REF175,REF258,SRPP117 and SRPP204 may form protein complexes and bind to RP.In addition to the members of the REF/SRPP family,20 proteins interacting with REF138 were mainly involved in the spliceosome and endocytic metabolic pathways,and 50 proteins interacting with REF258 were involved in the regulation of lipid metabolism and the synthesis of secondary metabolites.In conclusion,functional analysis of differential proteins and the interaction proteins responsive to ethylene stimulation on LRP and SRP might preliminarily reveal the metabolic regulatory mechanism of LRP and SRP in regulating natural rubber synthesis in response to exogenous ethylene stimulation.