Article(id=1276601105219977587, tenantId=1146029695717560320, journalId=1235980609244409860, issueId=1276600957765021779, articleNumber=null, orderNo=null, doi=10.3969/j.issn.1000-2561.2024.06.005, pmid=null, cstr=null, oa=null, hot=null, price=null, onlineType=0, articleFormat=0, articleType=null, articleTypeStr=null, receivedDate=1690819200000, receivedDateStr=2023-08-01, revisedDate=1694102400000, revisedDateStr=2023-09-08, acceptedDate=null, acceptedDateStr=null, onlineDate=1782295021585, onlineDateStr=2026-06-24, pubDate=1719244800000, pubDateStr=2024-06-25, doiRegisterDate=null, doiRegisterDateStr=null, onlineIssueDate=1782295021585, onlineIssueDateStr=2026-06-24, onlineJustAcceptDate=null, onlineJustAcceptDateStr=null, onlineFirstDate=null, onlineFirstDateStr=null, sourceXml=null, magXml=null, createTime=1782295021585, creator=13701087609, updateTime=1782295021585, updator=13701087609, issue=Issue{id=1276600957765021779, tenantId=1146029695717560320, journalId=1235980609244409860, year='2024', volume='45', issue='6', pageStart='1095', pageEnd='1302', issueExtLink='null', onlineDate='null', pubDate='1719244800000', pubDateStr='2024-06-25', beforeIssueId=null, nextIssueId=null, price=null, status=1, issueComplete=1, articleOrder=1, issueType=-1, specialIssue=null, createTime=1782294986430, creator='13701087609', updateTime=1782348406834, updator='13701087609', preIssue=null, nextIssue=null, articleTotal=null, ext={EN=IssueExt(id=1276825019267285043, tenantId=1146029695717560320, journalId=1235980609244409860, issueId=1276600957765021779, language=EN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=), CN=IssueExt(id=1276825019271479348, tenantId=1146029695717560320, journalId=1235980609244409860, issueId=1276600957765021779, language=CN, specialIssueTitle=, coverIllustrator=null, specialIssueEditor=, specialIssueAbout=)}, issueFiles=null, downloadFileDto=null}, startPage=1127, endPage=1138, ext={EN=ArticleExt(id=1276601105794597237, articleId=1276601105219977587, tenantId=1146029695717560320, journalId=1235980609244409860, language=EN, title=Role of Large and Small Rubber Particles of Hevea Brasiliensis in Ethylene Regulated Natural Rubber Synthesis, columnId=1236256430337085821, journalTitle=Chinese Journal of Tropical Crops, columnName=Omics & Biotechnology, runingTitle=null, highlight=null, articleAbstract=

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.

, authors=null, authorsList=Dan WANG, Bingqiang XU, Yong SUN, Cunzhi PENG, Lili CHANG, Zheng TONG, authorCompany=null, correspAuthors=Zheng TONG, authorNote=null, correspAuthorsNote=null, copyrightStatement=null, copyrightOwner=null, extLink=null, articleAbsUrl=null, sourceXml=null, magXml=null, pdfUrl=null, pdf=null, pdfFileSize=null, pdfExtLink=null, richHtmlUrl=null, mobilePdfUrl=null, reviewReport=null, pdfFirstPage=null, abstractGraph=null, abstractGraphContent=null, abstractVideo=null, citation=null, cebUrl=null, magXmlContent=null, mapNumber=null, fund=null), CN=ArticleExt(id=1276601109099708803, articleId=1276601105219977587, tenantId=1146029695717560320, journalId=1235980609244409860, language=CN, title=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, columnId=1236256430517440904, journalTitle=热带作物学报, columnName=组学与生物技术, runingTitle=null, highlight=null, articleAbstract=

巴西橡胶树(Hevea brasiliensis)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(14.7 kDa)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。

, authors=

王丹(1984—),女,硕士,助理研究员,研究方向:热带作物蛋白质组学。

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* 仝征(TONG Zheng),E-mail:
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王丹(1984—),女,硕士,助理研究员,研究方向:热带作物蛋白质组学。

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王丹(1984—),女,硕士,助理研究员,研究方向:热带作物蛋白质组学。

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Journal of Experimental Botany, 2017, 68(18): 5045-5055., articleTitle=Subcellular localization and interactions among rubber particle proteins from Hevea brasiliensis, refAbstract=null), Reference(id=1276824436833645222, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, doi=null, pmid=null, pmcid=null, year=2020, volume=25, issue=21, pageStart=5110, pageEnd=null, url=null, language=null, rfNumber=[28], rfOrder=27, authorNames=BAE S W, JUNG S, CHOI S C, KIM M Y, RYU S B, journalName=Molecules, refType=null, unstructuredReference=BAE S W, JUNG S, CHOI S C, KIM M Y, RYU S B. Lipid composition of latex and rubber particles in Hevea brasiliensis and Taraxacum kok-saghyz[J]. Molecules, 2020, 25(21): 5110., articleTitle=Lipid composition of latex and rubber particles in Hevea brasiliensis and Taraxacum kok-saghyz, refAbstract=null), Reference(id=1276824437039166119, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, doi=null, pmid=null, pmcid=null, year=2022, volume=12, issue=1, pageStart=3734, pageEnd=null, url=null, language=null, rfNumber=[29], rfOrder=28, authorNames=KUROIWA F, NISHINO A, MANDAL Y, HONZAWA M, SUENAGA-HIROMORI M, SUZUKI K, TAKANI Y, MIYAGI-INOUE Y, YAMAGUCHI H, YAMASHITA S, TAKAHASHI S, TOZAWA Y, journalName=Scientific Reports, refType=null, unstructuredReference=KUROIWA F, NISHINO A, MANDAL Y, HONZAWA M, SUENAGA-HIROMORI M, SUZUKI K, TAKANI Y, MIYAGI-INOUE Y, YAMAGUCHI H, YAMASHITA S, TAKAHASHI S, TOZAWA Y. Reconstitution of prenyltransferase activity on nanodiscs by components of the rubber synthesis machinery of the Para rubber tree and guayule[J]. 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Journal of Biological Chemistry, 1994, 269(12): 9278-9283., articleTitle=Hevein, a lectin-like protein from Hevea brasiliensis (rubber tree) is involved in the coagulation of latex, refAbstract=null), Reference(id=1276824437458596523, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, doi=null, pmid=null, pmcid=null, year=2003, volume=24, issue=4, pageStart=1, pageEnd=5, url=null, language=null, rfNumber=[33], rfOrder=32, authorNames=黄瑾, 校现周, journalName=热带作物学报, refType=null, unstructuredReference=黄瑾, 校现周. 乙烯利和乙烯刺激对橡胶树胶乳中几丁质酶活性和胶乳产量的影响[J]. 热带作物学报, 2003, 24(4): 1-5., articleTitle=乙烯利和乙烯刺激对橡胶树胶乳中几丁质酶活性和胶乳产量的影响, refAbstract=null), Reference(id=1276824437534093996, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, doi=null, pmid=null, pmcid=null, year=2003, volume=24, issue=4, pageStart=1, pageEnd=5, url=null, language=null, rfNumber=[33], rfOrder=33, authorNames=HUANG J, XIAO X Z, journalName=Chinese Journal Tropical Crops, refType=null, unstructuredReference=HUANG J, XIAO X Z. Effects of ethylene and ethylene stimulation on chitinase activity and latex yield of Hevea brasiliensis[J]. Chinese Journal Tropical Crops, 2003, 24(4): 1-5. (in Chinese), articleTitle=Effects of ethylene and ethylene stimulation on chitinase activity and latex yield of Hevea brasiliensis, refAbstract=null)], funds=[Fund(id=1276824431653679754, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, awardId=31500557, language=CN, fundingSource=国家自然科学基金项目(31500557), fundOrder=null, country=null)], companyList=[AuthorCompany(id=1276824415287505476, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, xref=1., ext=[AuthorCompanyExt(id=1276824415295894085, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, companyId=1276824415287505476, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=1.Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Haikou, Hainan 571101, China), AuthorCompanyExt(id=1276824415312671302, 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companyId=1276824415434306119, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=2.中国热带农业科学院海口实验站(热带果树研究所),海南海口 571101)]), AuthorCompany(id=1276824415727907403, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, xref=3., ext=[AuthorCompanyExt(id=1276824416059257420, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, companyId=1276824415727907403, language=EN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3.Rubber Research Institute, Chinese Academy of Tropical Agricultural Sciences, Danzhou, Hainan 571737, China), AuthorCompanyExt(id=1276824416076034637, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, companyId=1276824415727907403, language=CN, country=null, province=null, city=null, postcode=null, companyName=null, departmentName=null, remark=3.中国热带农业科学院橡胶研究所,海南儋州 571737)])], figs=[ArticleFig(id=1276824428268876407, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 1, caption=SEM observation of RP, LRP and SRP in H. brasiliensis latex treated with ethylene and ddH2O, figureFileSmall=54z5OZ3qVTN9+KtlUyCjcQ==, figureFileBig=XuUrTNQhVhyaVitpPMFbXw==, tableContent=null), ArticleFig(id=1276824428340179576, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图1, caption=乙烯和ddH2O处理后巴西橡胶树胶乳中RP、LRP和SRP的扫描电镜观察, figureFileSmall=54z5OZ3qVTN9+KtlUyCjcQ==, figureFileBig=XuUrTNQhVhyaVitpPMFbXw==, tableContent=null), ArticleFig(id=1276824428688306809, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 2, caption=Differential protein spots on 2D-DIGE gel of LRP and SRP proteins of rubber latex after ethylene treatment, figureFileSmall=Cq4zeWP/ftKpXlYzHghpGQ==, figureFileBig=fK4UUUWrSSKp7tSF1iJFeQ==, tableContent=null), ArticleFig(id=1276824428755415674, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图2, caption=乙烯处理后巴西橡胶树胶乳中LRP和SRP蛋白的2D-DIGE凝胶图谱, figureFileSmall=Cq4zeWP/ftKpXlYzHghpGQ==, figureFileBig=fK4UUUWrSSKp7tSF1iJFeQ==, tableContent=null), ArticleFig(id=1276824428822524539, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 3, caption=Enrichment of KEGG metabolic pathways of ethylene responsive differential proteins in LRP and SRP, figureFileSmall=6NtpSxaWtm+yHUE0gIk7ag==, figureFileBig=P57R/G/kRcRo5X4/srN7CQ==, tableContent=null), ArticleFig(id=1276824428902216316, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图3, caption=LRP和SRP中乙烯响应差异蛋白富集的KEGG代谢通路, figureFileSmall=6NtpSxaWtm+yHUE0gIk7ag==, figureFileBig=P57R/G/kRcRo5X4/srN7CQ==, tableContent=null), ArticleFig(id=1276824428981908093, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 4, caption=Western blot of key proteins among different components in H. brasiliensis latex

A: Western Blot of key proteins in different components; B: Western Blot of key proteins in LRP and SRP.

, figureFileSmall=bc2hElj/jKxnTJV2Vvqaiw==, figureFileBig=CKIQqQTFkza6IVq5biDXXQ==, tableContent=null), ArticleFig(id=1276824429065794174, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图4, caption=巴西橡胶树胶乳不同组分间关键蛋白质的Western Blot

A:不同组分中关键蛋白质的Western Blot;B:LRP和SRP中关键蛋白质的Western Blot。

, figureFileSmall=bc2hElj/jKxnTJV2Vvqaiw==, figureFileBig=CKIQqQTFkza6IVq5biDXXQ==, tableContent=null), ArticleFig(id=1276824429179040383, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 5, caption=Western blot of key proteins in H. brasiliensis C-serum and rubber particle between the ethylene treated groups, figureFileSmall=Ccf/6WJSulDx7S4dfSpRVw==, figureFileBig=gRPVhN/n9oX4eOat5GBDmg==, tableContent=null), ArticleFig(id=1276824430848373377, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图5, caption=巴西橡胶树胶C乳清和橡胶粒子中乙烯处理组关键蛋白质的Western Blot, figureFileSmall=Ccf/6WJSulDx7S4dfSpRVw==, figureFileBig=gRPVhN/n9oX4eOat5GBDmg==, tableContent=null), ArticleFig(id=1276824430949036674, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Fig. 6, caption=Enrichment of KEGG metabolic pathways of interacting proteins of REF138 and REF258, figureFileSmall=H/a5ehzWD8NZcNSpXOuk5g==, figureFileBig=ch5iAZs3Bl74m3El1GZKrw==, tableContent=null), ArticleFig(id=1276824431007756931, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=图6, caption=REF138和REF258互作蛋白富集的KEGG代谢通路, figureFileSmall=H/a5ehzWD8NZcNSpXOuk5g==, figureFileBig=ch5iAZs3Bl74m3El1GZKrw==, tableContent=null), ArticleFig(id=1276824431091643012, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Tab. 1, caption=

Differential protein information of LRP in latex of H. brasiliensis after ethylene and ddH2O treatment

, figureFileSmall=null, figureFileBig=null, tableContent=
编号No.差异比值(E/D)Average Ratio (E/D)差异可信性T-test蛋白名称Protein name蛋白登录号Protein accession理论等电点/理论分子量pI/MW (Da)质谱得分Score序列匹配率Coverage/%
L12.110.050Stromal 70 kDa heat shock-related protein, chloroplastic-likeXP_021650389.15.24/7522449912
L21.700.014Heat shock 70 kDa protein 5XP_021691174.15.36/7214964319
L33.880.004Malate dehydrogenaseXP_021640500.16.19/3617327928
L43.840.049Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524936440
L53.350.006Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524979663
L63.080.005Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961220426
L72.790.033Leucine aminopeptidase 1-likeXP_021672059.16.30/6164538210
L82.760.011Rubber elongation factor protein (REF138)XP_021653602.15.04/1471319125
L92.130.022Elongation factor 1-gammaXP_021660692.15.69/4868552329
L102.000.050Leucine aminopeptidase 1-likeXP_021672059.16.30/616452808
L111.950.018Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233165941
L121.840.012Phosphoglycerate kinase, cytosolicXP_021651095.15.90/4260064827
L131.690.015Protein DJ-1 homolog DXP_021645399.15.18/4298446918
L141.620.043Triosephosphate isomerase, cytosolicXP_021646026.15.90/2757172955
L151.610.013Patatin-like protein 2XP_021668042.15.13/4914282127
L161.600.032Actin-7XP_021666817.15.31/4189739728
L171.580.013Enolase 2XP_021644856.15.99/4799324317
L181.450.033Rubber elongation factor protein (REF138)XP_021653602.15.04/1471343138
L191.420.049Phosphoglycerate kinase, cytosolic-likeXP_021681983.15.67/4243875338
L201.380.044Isoflavone reductase-like proteinXP_021644235.15.50/3324038020
L211.300.036Cysteine proteinase inhibitor-likeXP_021677544.15.45/1123622934
L221.250.016Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233173974
L23–1.250.018Petacaspase-4-likeXP_021692568.15.01/4644313716
L24–1.260.017Phospholipase A1-IIdelta-likeXP_021650571.15.47/4741535817
L25–1.270.013Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961238426
L26–1.30.010Ricin B-like lectin EULS3XP_021658810.17.77/2072412012
L27–1.310.019Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529952230
L28–1.320.018Alpha-soluble NSF attachment protein 2-likeXP_021668979.15.12/3280218919
L29–1.340.020Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529920825
L30–1.410.047Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529922816
L31–1.410.027NADPH-dependent aldehyde reductase 1, chloroplasticXP_021670828.15.29/3230940424
L32–1.490.013Actin-7XP_021666817.15.31/4189746725
L33–1.530.042Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529955530
L34–1.590.043Uncharacterized protein LOC110668061 isoformX1XP_021684781.17.67/2950023216
L35–1.630.011Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529956030
L36–1.670.016Rubber elongation factor protein (REF138)XP_021653602.15.04/1471362955
L37–2.020.008Rubber elongation factor protein (REF138)XP_021653602.15.04/1471354855
), ArticleFig(id=1276824431183917701, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=表1, caption=

乙烯处理后巴西橡胶树胶乳中LRP差异蛋白信息

, figureFileSmall=null, figureFileBig=null, tableContent=
编号No.差异比值(E/D)Average Ratio (E/D)差异可信性T-test蛋白名称Protein name蛋白登录号Protein accession理论等电点/理论分子量pI/MW (Da)质谱得分Score序列匹配率Coverage/%
L12.110.050Stromal 70 kDa heat shock-related protein, chloroplastic-likeXP_021650389.15.24/7522449912
L21.700.014Heat shock 70 kDa protein 5XP_021691174.15.36/7214964319
L33.880.004Malate dehydrogenaseXP_021640500.16.19/3617327928
L43.840.049Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524936440
L53.350.006Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524979663
L63.080.005Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961220426
L72.790.033Leucine aminopeptidase 1-likeXP_021672059.16.30/6164538210
L82.760.011Rubber elongation factor protein (REF138)XP_021653602.15.04/1471319125
L92.130.022Elongation factor 1-gammaXP_021660692.15.69/4868552329
L102.000.050Leucine aminopeptidase 1-likeXP_021672059.16.30/616452808
L111.950.018Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233165941
L121.840.012Phosphoglycerate kinase, cytosolicXP_021651095.15.90/4260064827
L131.690.015Protein DJ-1 homolog DXP_021645399.15.18/4298446918
L141.620.043Triosephosphate isomerase, cytosolicXP_021646026.15.90/2757172955
L151.610.013Patatin-like protein 2XP_021668042.15.13/4914282127
L161.600.032Actin-7XP_021666817.15.31/4189739728
L171.580.013Enolase 2XP_021644856.15.99/4799324317
L181.450.033Rubber elongation factor protein (REF138)XP_021653602.15.04/1471343138
L191.420.049Phosphoglycerate kinase, cytosolic-likeXP_021681983.15.67/4243875338
L201.380.044Isoflavone reductase-like proteinXP_021644235.15.50/3324038020
L211.300.036Cysteine proteinase inhibitor-likeXP_021677544.15.45/1123622934
L221.250.016Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233173974
L23–1.250.018Petacaspase-4-likeXP_021692568.15.01/4644313716
L24–1.260.017Phospholipase A1-IIdelta-likeXP_021650571.15.47/4741535817
L25–1.270.013Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961238426
L26–1.30.010Ricin B-like lectin EULS3XP_021658810.17.77/2072412012
L27–1.310.019Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529952230
L28–1.320.018Alpha-soluble NSF attachment protein 2-likeXP_021668979.15.12/3280218919
L29–1.340.020Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529920825
L30–1.410.047Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529922816
L31–1.410.027NADPH-dependent aldehyde reductase 1, chloroplasticXP_021670828.15.29/3230940424
L32–1.490.013Actin-7XP_021666817.15.31/4189746725
L33–1.530.042Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529955530
L34–1.590.043Uncharacterized protein LOC110668061 isoformX1XP_021684781.17.67/2950023216
L35–1.630.011Rubber elongation factor protein-like isoform X1(REF258)XP_021653592.15.21/2529956030
L36–1.670.016Rubber elongation factor protein (REF138)XP_021653602.15.04/1471362955
L37–2.020.008Rubber elongation factor protein (REF138)XP_021653602.15.04/1471354855
), ArticleFig(id=1276824431271998086, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Tab. 2, caption=

Differential protein information of SRP in latex of H. brasiliensis after ethylene and ddH2O treatment

, figureFileSmall=null, figureFileBig=null, tableContent=
编号No.差异比值(E/D)Average ratio (E/D)差异可信性T-test蛋白名称Protein name蛋白登录号Protein accession理论等电点/理论分子量pI/MW (Da)质谱得分Score序列匹配率Coverage/%
S14.250.050Rubber elongation factor protein (REF138)XP_021653602.15.04/1471323725
S24.191E-04Rubber elongation factor protein (REF138)XP_021653602.15.04/1471334438
S34.160.001Rubber elongation factor protein (REF138)XP_021653602.15.04/1471338455
S43.760.002Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328638
S52.199E-05Small rubber particle protein isoform X1(SRPP204)XP_021653597.1|4.80/2233137260
S61.980.047Nuclear transport factor 2XP_021640386.15.65/1369211726
S71.910.010Pro-heveinXP_021650927.15.89/2315410523
S81.900.019Pro-heveinXP_021650927.15.89/2315412023
S91.780.010Pro-heveinXP_021650927.15.89/2315411114
S101.720.046Rubber elongation factor protein (REF138)XP_021653602.15.04/1471317525
S111.666E-04Rubber elongation factor protein (REF138)XP_021653602.15.04/1471324038
S121.620.010Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328338
S131.610.021Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328242
S141.600.050Thioredoxin-like protein ClotXP_021656682.15.69/1506014046
S151.590.011Actin-depolymerizing factor 2XP_021663200.15.92/1622713123
S161.580.002Small rubber particle protein-like isoform X1(SRPP117)XP_021653603.14.61/1276916861
S171.570.01317.1 kDa class II heat shock protein-likeXP_021661608.15.52/1764820335
S181.500.01517.9 kDa class II heat shock protein-likeXP_021655193.15.80/1753513219
S191.480.009Actin-depolymerizing factor 2-likeXP_021689182.15.92/1622014651
S201.470.02117.3 kDa class I heat shock protein-likeXP_021638116.16.00/1833555143
S211.420.010Putative 4-hydroxy-4-methyl-2-oxoglutaratealdolase 2XP_021673665.16.13/1799825730
S221.400.049Eukaryotic translation initiation factor 5A-likeXP_021645712.15.60/1768931135
S231.390.015Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524919845
S241.390.047Peroxiredoxin-2B-likeXP_021675296.15.55/175429820
S251.380.01617.3 kDa class I heat shock protein-likeXP_021638116.16.00/1833532725
S261.340.046Probable phospholipid hydroperoxide glutathione peroxidaseXP_021674982.18.78/26698419
S271.300.050ATP synthase CF1 beta subunit (chloroplast)YP_004327669.15.17/53665377
S281.250.039Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961218321
S291.240.010Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233144860
S301.210.016Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233143660
S311.200.046Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233127157
S32–1.200.049Probable glutathione S-transferaseXP_021670613.16.23/25563998
S33–1.230.018Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529942134
S34–1.230.020Uncharacterized protein LOC110633924XP_021638455.18.91/318999913
S35–1.250.023Triosephosphate isomerase, cytosolic-likeXP_021677209.15.76/2753319122
S36–1.250.041Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529917212
S37–1.280.047Triosephosphate isomerase, cytosolicXP_021646026.15.90/2757120817
S38–1.290.008Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529917423
S39–1.310.050Uncharacterized protein LOC110659451XP_021673087.16.09/3159032324
S40–1.340.005Rubber elongation factor protein (REF138)XP_021653602.15.04/1471318925
S41–1.350.033Rubber elongation factor protein (REF138)XP_021653602.15.04/1471316525
S42–1.350.039Rubber elongation factor protein (REF138)XP_021653602.15.04/1471329025
S43–1.380.035Rran-binding protein 1 homolog b-likeXP_021645721.14.85/2541930031
S44–1.380.040Rubber elongation factor protein (REF138)XP_021653602.15.04/1471317342
S45–1.420.017Ricin B-like lectin R40G3XP_021663961.16.16/3407920917
S46–1.420.050Rubber elongation factor protein (REF138)XP_021653602.15.04/1471332655
S47–1.450.023Ricin B-like lectin R40G3XP_021663961.16.16/3407916517
S48–1.460.047Guanosine nucleotide diphosphate dissociation inhibitor 2XP_021673535.15.54/5014214311
S49–1.470.040Elongation factor 1-delta 1-like isoform X1XP_021690229.14.47/2540825218
S50–1.490.020Patatin-like protein 2XP_021672059.15.13/4914214214
S51–1.60.023Protein EARLY-RESPONSIVE TODEHYDRATION 7, chloroplastic-likeXP_021658906.15.11/4876131421
S52–1.630.046Probable mediator of RNA polymerase II transcription subunit 37c isoform X1XP_021663050.15.14/7163939612
S53–1.730.002Heat shock 70 kDa protein 5XP_021691174.15.36/7214931919
S54–1.920.030Leucine aminopeptidase 1-likeXP_021672059.16.30/616452169
S55–2.170.038Phospholipase A1-IIdelta-likeXP_021650571.15.47/4741550818
S56–2.530.041Eukaryotic initiation factor 4A-11XP_021638642.15.49/4709547035
), ArticleFig(id=1276824431355884167, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=表2, caption=

乙烯和ddH2O处理后巴西橡胶树胶乳中SRP差异蛋白信息

, figureFileSmall=null, figureFileBig=null, tableContent=
编号No.差异比值(E/D)Average ratio (E/D)差异可信性T-test蛋白名称Protein name蛋白登录号Protein accession理论等电点/理论分子量pI/MW (Da)质谱得分Score序列匹配率Coverage/%
S14.250.050Rubber elongation factor protein (REF138)XP_021653602.15.04/1471323725
S24.191E-04Rubber elongation factor protein (REF138)XP_021653602.15.04/1471334438
S34.160.001Rubber elongation factor protein (REF138)XP_021653602.15.04/1471338455
S43.760.002Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328638
S52.199E-05Small rubber particle protein isoform X1(SRPP204)XP_021653597.1|4.80/2233137260
S61.980.047Nuclear transport factor 2XP_021640386.15.65/1369211726
S71.910.010Pro-heveinXP_021650927.15.89/2315410523
S81.900.019Pro-heveinXP_021650927.15.89/2315412023
S91.780.010Pro-heveinXP_021650927.15.89/2315411114
S101.720.046Rubber elongation factor protein (REF138)XP_021653602.15.04/1471317525
S111.666E-04Rubber elongation factor protein (REF138)XP_021653602.15.04/1471324038
S121.620.010Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328338
S131.610.021Rubber elongation factor protein (REF138)XP_021653602.15.04/1471328242
S141.600.050Thioredoxin-like protein ClotXP_021656682.15.69/1506014046
S151.590.011Actin-depolymerizing factor 2XP_021663200.15.92/1622713123
S161.580.002Small rubber particle protein-like isoform X1(SRPP117)XP_021653603.14.61/1276916861
S171.570.01317.1 kDa class II heat shock protein-likeXP_021661608.15.52/1764820335
S181.500.01517.9 kDa class II heat shock protein-likeXP_021655193.15.80/1753513219
S191.480.009Actin-depolymerizing factor 2-likeXP_021689182.15.92/1622014651
S201.470.02117.3 kDa class I heat shock protein-likeXP_021638116.16.00/1833555143
S211.420.010Putative 4-hydroxy-4-methyl-2-oxoglutaratealdolase 2XP_021673665.16.13/1799825730
S221.400.049Eukaryotic translation initiation factor 5A-likeXP_021645712.15.60/1768931135
S231.390.015Elicitor-responsive protein 3-like isoform X1XP_021663495.14.06/1524919845
S241.390.047Peroxiredoxin-2B-likeXP_021675296.15.55/175429820
S251.380.01617.3 kDa class I heat shock protein-likeXP_021638116.16.00/1833532725
S261.340.046Probable phospholipid hydroperoxide glutathione peroxidaseXP_021674982.18.78/26698419
S271.300.050ATP synthase CF1 beta subunit (chloroplast)YP_004327669.15.17/53665377
S281.250.039Rubber elongation factor protein-like (REF175)XP_021653600.15.28/1961218321
S291.240.010Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233144860
S301.210.016Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233143660
S311.200.046Small rubber particle protein isoform X1(SRPP204)XP_021653597.14.80/2233127157
S32–1.200.049Probable glutathione S-transferaseXP_021670613.16.23/25563998
S33–1.230.018Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529942134
S34–1.230.020Uncharacterized protein LOC110633924XP_021638455.18.91/318999913
S35–1.250.023Triosephosphate isomerase, cytosolic-likeXP_021677209.15.76/2753319122
S36–1.250.041Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529917212
S37–1.280.047Triosephosphate isomerase, cytosolicXP_021646026.15.90/2757120817
S38–1.290.008Rubber elongation factor protein-like isoformX1 (REF258)XP_021653592.15.21/2529917423
S39–1.310.050Uncharacterized protein LOC110659451XP_021673087.16.09/3159032324
S40–1.340.005Rubber elongation factor protein (REF138)XP_021653602.15.04/1471318925
S41–1.350.033Rubber elongation factor protein (REF138)XP_021653602.15.04/1471316525
S42–1.350.039Rubber elongation factor protein (REF138)XP_021653602.15.04/1471329025
S43–1.380.035Rran-binding protein 1 homolog b-likeXP_021645721.14.85/2541930031
S44–1.380.040Rubber elongation factor protein (REF138)XP_021653602.15.04/1471317342
S45–1.420.017Ricin B-like lectin R40G3XP_021663961.16.16/3407920917
S46–1.420.050Rubber elongation factor protein (REF138)XP_021653602.15.04/1471332655
S47–1.450.023Ricin B-like lectin R40G3XP_021663961.16.16/3407916517
S48–1.460.047Guanosine nucleotide diphosphate dissociation inhibitor 2XP_021673535.15.54/5014214311
S49–1.470.040Elongation factor 1-delta 1-like isoform X1XP_021690229.14.47/2540825218
S50–1.490.020Patatin-like protein 2XP_021672059.15.13/4914214214
S51–1.60.023Protein EARLY-RESPONSIVE TODEHYDRATION 7, chloroplastic-likeXP_021658906.15.11/4876131421
S52–1.630.046Probable mediator of RNA polymerase II transcription subunit 37c isoform X1XP_021663050.15.14/7163939612
S53–1.730.002Heat shock 70 kDa protein 5XP_021691174.15.36/7214931919
S54–1.920.030Leucine aminopeptidase 1-likeXP_021672059.16.30/616452169
S55–2.170.038Phospholipase A1-IIdelta-likeXP_021650571.15.47/4741550818
S56–2.530.041Eukaryotic initiation factor 4A-11XP_021638642.15.49/4709547035
), ArticleFig(id=1276824431439770248, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=EN, label=Tab. 3, caption=

Information of members of the REF/SRPP protein family with interaction to REF138 and REF258

, figureFileSmall=null, figureFileBig=null, tableContent=
DAP蛋白登录号Protein accession蛋白名称Protein name序列匹配率Coverage/%匹配多肽数Peptides
REF138XP_021653597.1Small rubber particle protein isoform X1 (SRPP204)71.611
XP_021653602.1Rubber elongation factor protein (REF138)54.36
XP_021653600.1Rubber elongation factor protein-like (REF175)21.74
XP_021653603.1Small rubber particle protein-like isoform X1 (SRPP117)28.22
XP_021653604.1Small rubber particle protein-like isoform X225.72
REF258XP_021653597.1Small rubber particle protein isoform X1 (SRPP204)71.611
XP_021653602.1Rubber elongation factor protein (REF138)57.28
XP_021653592.1Rubber elongation factor protein-like isoform X1 (REF258)38.47
XP_021653600.1Rubber elongation factor protein-like (REF175)12.62
XP_021653603.1Small rubber particle protein-like isoform X1 (SRPP117)28.22
), ArticleFig(id=1276824431519462025, tenantId=1146029695717560320, journalId=1235980609244409860, articleId=1276601105219977587, language=CN, label=表3, caption=

与REF138和REF258存在相互作用关系的REF/SRPP蛋白家族成员信息

, figureFileSmall=null, figureFileBig=null, tableContent=
DAP蛋白登录号Protein accession蛋白名称Protein name序列匹配率Coverage/%匹配多肽数Peptides
REF138XP_021653597.1Small rubber particle protein isoform X1 (SRPP204)71.611
XP_021653602.1Rubber elongation factor protein (REF138)54.36
XP_021653600.1Rubber elongation factor protein-like (REF175)21.74
XP_021653603.1Small rubber particle protein-like isoform X1 (SRPP117)28.22
XP_021653604.1Small rubber particle protein-like isoform X225.72
REF258XP_021653597.1Small rubber particle protein isoform X1 (SRPP204)71.611
XP_021653602.1Rubber elongation factor protein (REF138)57.28
XP_021653592.1Rubber elongation factor protein-like isoform X1 (REF258)38.47
XP_021653600.1Rubber elongation factor protein-like (REF175)12.62
XP_021653603.1Small rubber particle protein-like isoform X1 (SRPP117)28.22
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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
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王丹 1 , 徐兵强 2 , 孙勇 3 , 彭存智 1 , 常丽丽 1 , 仝征 1, *
热带作物学报 | 组学与生物技术 2024,45(6): 1127-1138
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热带作物学报 |组学与生物技术 2024 , 45 (6) : 1127 -1138
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
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王丹1, 徐兵强2, 孙勇3, 彭存智1, 常丽丽1, 仝征1, *
作者信息
  • 1.中国热带农业科学院热带生物技术研究所,海南海口 571101
  • 2.中国热带农业科学院海口实验站(热带果树研究所),海南海口 571101
  • 3.中国热带农业科学院橡胶研究所,海南儋州 571737
通讯作者:
* 仝征(TONG Zheng),E-mail:
Role of Large and Small Rubber Particles of Hevea Brasiliensis in Ethylene Regulated Natural Rubber Synthesis
Dan WANG1, Bingqiang XU2, Yong SUN3, Cunzhi PENG1, Lili CHANG1, Zheng TONG1, *
Affiliations
  • 1.Institute of Tropical Bioscience and Biotechnology, Chinese Academy of Tropical Agricultural Sciences, Haikou, Hainan 571101, China
  • 2.Haikou Experimental Station (Institute of Tropical Fruit Tree Research), Chinese Academy of Tropical Agricultural Sciences, Haikou, Hainan 571101, China
  • 3.Rubber Research Institute, Chinese Academy of Tropical Agricultural Sciences, Danzhou, Hainan 571737, China
出版时间: 2024-06-25 doi: 10.3969/j.issn.1000-2561.2024.06.005
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巴西橡胶树(Hevea brasiliensis)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(14.7 kDa)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。

巴西橡胶树  /  橡胶粒子  /  乙烯  /  天然橡胶合成调控  /  蛋白互作

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.

Hevea brasiliensis  /  rubber particles  /  ethylene  /  regulation of natural rubber biosynthesis  /  protein interaction
王丹, 徐兵强, 孙勇, 彭存智, 常丽丽, 仝征. 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. 热带作物学报, 2024 , 45 (6) : 1127 -1138 . DOI: 10.3969/j.issn.1000-2561.2024.06.005
Dan WANG, Bingqiang XU, Yong SUN, Cunzhi PENG, Lili CHANG, Zheng TONG. Role of Large and Small Rubber Particles of Hevea Brasiliensis in Ethylene Regulated Natural Rubber Synthesis[J]. Chinese Journal of Tropical Crops, 2024 , 45 (6) : 1127 -1138 . DOI: 10.3969/j.issn.1000-2561.2024.06.005
巴西橡胶树(Hevea brasiliensis)是合成天然橡胶的重要产胶植物。位于其形成层中的乳管是合成天然橡胶的重要部位。乳管细胞中的橡胶粒子(RP)是天然橡胶合成的关键细胞器,内部储存天然橡胶,外部由含有蛋白质和其他成分的单层膜组织包被[1]。RP的直径通常在0.08~4 μm之间[2],不同直径的RP合成天然橡胶的效率不同,小橡胶粒子(SRP,直径小于0.35 μm)中天然橡胶合成效率高于大橡胶粒子(LRP,直径大于0.75 μm)[2-3]。结合在RP膜上的蛋白质分别参与天然橡胶生物合成、应激或防御相关反应、蛋白质加工和折叠、信号转导和细胞转运等代谢调控[4]
天然橡胶生物合成过程中,蔗糖作为底物经酶促反应生成乙酰辅酶A[5],经由甲羟戊酸途径(MVA)生成橡胶单体焦磷酸异戊烯基(IPP)。在多种酶的共同催化作用下,经起始、聚合及终止等步骤合成天然橡胶[6-9]。其中橡胶链的聚合在RP上完成,由多种蛋白酶类共同催化调控[8]。橡胶延伸因子蛋白(REF)、小橡胶粒子蛋白(SRPP)和顺式异戊烯基转移酶(CPT)是RP中参与橡胶生物合成的关键蛋白质[4]。REF的二级结构主要由β-折叠构成且高度疏水,以嵌入的方式结合在RP膜上[10],有利于调控跨膜运输过程[11]。胶乳中LRP的数量占RP总数量的6%以下,但其中的天然橡胶含量超过RP中总含量的93%[12],LRP中存储更多的天然橡胶。SRPP的二级结构主要由α-螺旋构成,仅与RP膜脂质表面结合[10],体外实验表明SRPP在橡胶链聚合过程中促进了IPP的掺入[13],并且能够提升橡胶粒子稳定性[14]。3-羟基-3-甲基戊二酰辅酶A合成酶(HMGS)是MVA中的关键酶,在SRP上积累较多[3]。SRP与天然橡胶合成的起始过程相关,LRP与橡胶链合成的终止过程相关。然而,LRP和SRP分别调控天然橡胶生物合成的作用机制仍不清楚。
研究发现外源乙烯可以提高巴西橡胶树天然橡胶产量。施加外源乙烯后,能够抑制胶乳中参与凝集过程的一系列酶和蛋白质表达,延迟乳管末端堵塞过程[15],延长胶乳流出时间从而获得更多的天然橡胶。RP及其蛋白质与胶乳凝集密切相关。橡胶树胶乳中的凝集素蛋白识别RP蛋白并诱导RP的迅速聚集[16]。然而,RP蛋白响应乙烯信号并抑制胶乳凝集的调节机制仍不清楚。另一方面,外源乙烯的施用可通过增加RP上天然橡胶合成关键蛋白质的积累促进其合成过程[17],其中REF/SRPP蛋白质家族在此过程中可能发挥重要作用[18],但具体的调控机制还需进一步研究。
因此,本研究关注巴西橡胶树胶乳中不同大小的橡胶粒子蛋白分别在乙烯刺激过程中的蛋白积累差异,并分析天然橡胶合成代谢调控关键蛋白的互作蛋白功能,明确LRP、SRP上响应乙烯刺激的代谢调控机制,进一步明确LRP、SRP在天然橡胶合成过程中的作用。
供试材料为未进行过割胶处理的10年树龄巴西橡胶树(H. brasiliensis Mull. Arg.,R7-33-97)。处理方法参照WANG等[19]的方法,在距离地面120 cm处的树干上开割线,24 h后在割线处涂抹5 mL的3%(V/V)乙烯刺激剂作为处理组(E),使用等体积的ddH2O涂抹割线作为对照组(D),使用封口膜对处理后的割线进行包裹。处理24 h后进行割胶,在冰上收集流出的胶乳。
冰上收集的胶乳在40 000´g,4 ℃条件下离心60 min,收集离心管上层的RP,按照1∶10(w/V)的比例用预冷的洗脱溶液(WS,20 mmol/L Tris-HCl,300 mmol/L甘露醇,0.5 mmol/L DTT,pH 7.0)洗涤。使用基于percoll介质的密度梯度离心法(percoll,1.31 mg/mL,Pharmacia,密度梯度由上至下依次为5%、15%、30%,V/V)将洗涤后的RP分离为LRP和SRP[20]。具体操作如下:将RP按照1∶10(w/V)的比例重新悬浮在含有30%(V/V)percoll的预冷WS中,转移到离心管底层。加入含有15%(V/V)percoll的预冷WS作为中间层和5%(V/V)percoll的预冷WS作为顶层。在5000´g,4 ℃条件下离心15 min。离心后顶层样本为LRP,底层样本为SRP。
将RP、LRP和SRP悬浮在固定溶液(50 mmol/L二甲基砷钠,1%(w/V)单宁酸,6%(V/V)戊二醛)中室温静置2 h。离心收集固定后的RP、LRP和SRP,ddH2O洗涤3次。加入1%(w/V)的四氧化锇室温固定1 h,ddH2O洗涤2次后加入适量的ddH2O重新悬浮。将悬浮液滴加在锡箔纸上,风干后的RP、LRP和SRP样品经扫描电子显微镜(SEM)观测。
使用酚抽提法获得LRP和SRP的蛋白质样本。加入双倍LRP和SRP体积的蛋白提取液BPP(100 mmol/L EDTA,50 mmol/L硼砂,50 mmol/L维生素C,1%(w/V)PVPP,1%(w/V)Triton X-100,2%(w/V)β-巯基乙醇,30%(w/V)蔗糖,pH 8.0),震荡10 min后加入等体积的Tris饱和苯酚(pH>7.8),混合物在15 000´g,4 ℃条件下离心5 min。取离心后的上层液体再加入等体积BPP,震荡5 min后相同条件离心,取离心后的上层液体加入5倍体积的预冷硫酸铵饱和甲醇溶液并在–20 ℃环境中静置12 h以上,在15 000´g,4 ℃条件下离心15 min获得粗蛋白,加入缓冲液LB [7 mol/L尿素,2 mol/L硫脲,30 mmol/L Tris,2%(w/V)CHAPS,pH 8.5]充分溶解获得蛋白样本,以BSA为标准,通过Bradford法测定样本浓度。
将获得的蛋白质样本进行双向荧光差异凝胶电泳(DIGE)。50 μg SRP蛋白样本和LRP蛋白样本,分别以400 pmol/L的Cy3和Cy5荧光染料标记,所有样本等量混合后用Cy2荧光染料标记作为内标。电泳后凝胶图像使用Typhoon Trio扫描仪(GE healthcare)采集,DIGE凝胶图谱经DeCyder 7.0软件分析获得差异蛋白(differential accumulation proteins,DAPs),差异蛋白经胰蛋白酶酶解后进行质谱(MALDI-TOF/TOF MS,AB-sciex)分析检测。
胶乳(TL)在40 000´g,4 ℃条件下离心60 min,离心后最上层是RP,中间层是C乳清(CS),底部是黄色体(LO)。TL、CS、LO蛋白样本按照1.2.2中的方法提取。蛋白样本完成单向凝胶电泳,经膜转印后,使用5%(w/V)脱脂牛奶封闭,分别与实验室前期获得的多个DAPs多克隆兔血清抗体和HRP缀合的羊抗兔IgG孵育,经显色后由LAS 4000 mini成像仪(GE Healthcare)采集图谱。
采用免疫沉淀结合液质联用质谱技术,对巴西橡胶树胶乳中关键DAPs的互作蛋白进行鉴定。使用多克隆兔血清抗体及Protein A+G Agarose(Fast Flow,for IP)试剂盒(Beyotime)进行免疫沉淀实验。具体步骤如下:冰上采集1 mL的新鲜胶乳加入等体积预冷的WS溶液,加入2 μg多克隆兔血清抗体后4 ℃孵育过夜。加入100 μL充分重悬的Protein A+G Agarose,4 ℃孵育3 h后在1000´g,4 ℃条件下离心5 min,除去上清后加入1 mL的PBS溶液洗涤沉淀5次,加入适量缓冲液LB获得互作蛋白样本。对该样本进行胰蛋白酶解后,使用液质联用质谱HPLC(Dionex Ultra 3000 nano)-ESI-MS/MS(TripleTOF 5600,ABsciex)进行鉴定,经蛋白质数据库分析后,获得互作蛋白质信息。
采用基于percoll的密度梯度离心法,分别从乙烯和ddH2O处理后的巴西橡胶树胶乳中成功分离获得了高纯度的LRP和SRP。通过SEM观测到RP的直径范围在0.05 μm至1.0 μm(图1)。直径小于0.3 μm的为SRP,同时直径大于0.5 μm的为LRP。
通过2D-DIGE对乙烯处理(E)和ddH2O处理(D)24 h后的橡胶树胶乳LRP和SRP蛋白质进行差异分析(图2)。在LRP中,共检测到37个蛋白点在处理组和对照组之间发生变化(P<0.01,T<0.05,变化率高于1.2倍)(图2表1)。从这些蛋白点中鉴定到25个差异蛋白质(DAPs)。其中,13个蛋白点对应的4个DAPs与天然橡胶合成直接相关,属于REF/SRPP家族成员。蛋白点L6和L25鉴定到同一种含有175个氨基酸的橡胶延伸因子蛋白(REF175),在乙烯刺激后2种分子量相同但等电点不同的REF175蛋白亚型的含量在LRP上呈现不同的变化趋势,蛋白点L6高调3.08倍,蛋白点L25低调1.27倍。L8、L18、L36和L37均鉴定为含有138个氨基酸的橡胶伸长因子蛋白(REF138),这4种不同蛋白亚型的积累量在乙烯刺激后具有不同变化趋势,等电点较高的REF138亚型经乙烯刺激后积累量增加;等电点较低的REF138亚型积累量减少。L27、L29、L30、L33和L35均鉴定为含有258个氨基酸的橡胶伸长因子蛋白(REF258),这5种不同蛋白亚型的积累量经乙烯刺激后在LRP上的积累量都明显减少。S11和S22鉴定为含有204个氨基酸的小橡胶颗粒蛋白(SRPP204),这2个蛋白亚型经乙烯刺激后在LRP上积累量均呈现增加的趋势。除REF/SRPP家族成员外还鉴定到21个DAPs,分别参与丙酮酸代谢、内吞、三羧酸循环、谷胱甘肽代谢、RNA降解、半胱氨酸和蛋氨酸代谢、磷酸肌醇代谢等通路(图3表1)。
SRP中,响应乙烯刺激积累量变化的蛋白点较LRP多,共检测到56个蛋白点在对照组和处理组之间发生变化(图2表2)。其中22个蛋白点鉴定到5个REF/SRPP家族成员。在乙烯刺激后,REF175(S28)在SRP上的积累量增加。有13个蛋白点鉴定为REF138,其中8个响应乙烯刺激积累量增加。REF258的3个蛋白亚型(S33、S36和S38)受乙烯刺激后在SRP上的积累量减少(图2表2)。SRPP204的4个蛋白亚型(S5、S29、S30和S31)经乙烯刺激后在SRP上积累量均增加。一种含有117个氨基酸的小橡胶颗粒蛋白亚型(SRPP117,S16)受乙烯刺激后SRP上的积累量增加。此外,鉴定到29个不属于REF/SRPP家族成员的DAPs,分别参与内质网中的蛋白质加工、谷胱甘肽代谢、糖酵解/糖异生、花生四烯酸代谢、剪接体和氧化磷酸化等代谢通路(图3表2)。
为了检测与天然橡胶合成调控相关关键蛋白在胶乳中不同组分间的分布差异,对TL、CS、LO、RP、LRP和SRP等不同胶乳组分蛋白样本进行Western Blot分析。结果发现,响应乙烯差异积累的REF/SRPP家族成员REF138、REF175和REF258主要分布在RP上,并且蛋白积累量在LRP上要多于SRP。REF258在胶乳不同组分中都检测到分子量不同的2条带,这2种REF258主要在RP中积累,CS中检测到少许,LO中未检出。SRPP117主要在CS中大量积累,RP中有微量检出,并主要在SRP中积累(图4)。SRPP117与其他具有完整REF domain的REF/SRPP蛋白家族成员不同,其REF domain存在部分缺失;并且与SRPP亚家族成员具有短N-末端及可变长度的C端不同,只有一个位于REF结构域两侧的短N端和C端序列[18]。Western Blot检测到SRPP117在CS和SRP组分中积累。SRPP117的不同蛋白亚型在SRP上响应乙烯刺激积累量增加(图2表2),并存在与REF138和REF258相互作用的可能(表3),SRPP117可能通过参与REF138和REF258构成的膜蛋白复合体参与天然橡胶合成的调控。
以ddH2O处理组作为对照,比较乙烯刺激后CS、RP、LRP及SRP上REF/SRPP家族蛋白的积累差异。Western Blot分析显示,乙烯刺激后SRPP117仅在CS组分中检出,并响应乙烯刺激积累量明显增加;REF138在CS及RP组分中都略有增加;REF175在RP中且主要是LRP中积累量增加;REF258在RP及SRP中变化不明显(图5)。研究认为REF是LRP上的主要蛋白而SRPP是SRP上的主要蛋白[10-1113],但有一项针对LRP与SRP间的差异蛋白研究表明,REF138在LRP和SRP上的表达量接近[3],本研究也证实这一发现,并且REF138在LRP和SRP中均响应乙烯刺激积累量增加(图5)。经2D-DIGE证实LRP及SRP中都存在不同等电点的REF138蛋白亚型,并且这些蛋白亚型在乙烯刺激后的积累量存在差异(图2表1表2)。在LRP上低于标准等电点(4.80)的REF138蛋白亚型响应乙烯刺激低积累,高于标准等电点的REF138蛋白亚型响应乙烯刺激高积累(表1)。与LRP不同,SRP上有更多的REF138蛋白亚型响应乙烯刺激发生变化,并且具有标准分子量(14.7 kDa)的不同蛋白亚型绝大部分响应乙烯刺激上调,高于标准分子量的蛋白亚型响应乙烯刺激下调(表2),暗示了乙烯可能通过诱导REF138不同蛋白亚型的差异积累调控天然橡胶合成。2D-DIGE发现LRP和SRP上REF258的一些不同等电点的蛋白亚型表现出明显下调趋势(表1表2),但Western Blot分析发现REF258的总含量变化不大(图5),这些结果暗示乙烯刺激后REF258可能通过调节其不同蛋白亚型等电点的改变对天然橡胶合成过程进行调控,具体作用机制需进一步研究加以证实。
对巴西橡胶树胶乳中与REF138和REF258互作的蛋白分别进行鉴定后发现,与REF138存在相互作用关系的可信蛋白有25个,其中与已知的天然橡胶合成调控相关的蛋白有5个;与REF258存在相互作用关系的可信蛋白有55个,其中与已知的天然橡胶合成调控相关的蛋白有5个(表3)。通过对比发现,REF138、REF175、REF258、SRPP117与SRPP204等5个天然橡胶合成调控关键蛋白质间存在相互作用,可能组成蛋白复合体存在于RP膜上,进而高效调控天然橡胶的合成。
除与天然橡胶合成直接相关的蛋白外,与REF138互作的蛋白还参与到乙醛酸和二羧酸代谢、RNA降解与转运、内吞作用以及内质网中的蛋白质加工等过程;与REF258互作的蛋白参与到甘油磷脂代谢、戊糖磷酸途径、糖酵解/糖异生、异喹啉生物碱生物合成、脂肪酸代谢、RNA降解、氧化磷酸化及MAPK信号通路等代谢过程中(图6)。此外,REF138和REF258都能够与一种几丁质酶hevamine-A相互作用,可能参与了胶乳凝集过程的调控。
乙烯响应因子蛋白(ethylene response factor)是茉莉酸和乙烯信号通路的重要因子,调控植物防御相关基因协同作用[21]。巴西橡胶树中的乙烯响应因子蛋白(HbERF-IXc5)在乳管代谢和应激耐受过程中发挥功能,促进乳胶的再生[22]。HbERF-IXc4在胶乳中有较高的转录丰度,乙烯刺激后激活其靶基因HbSUT3促进其表达[23]HbSUT3的高水平表达能够促进天然橡胶产量。本研究中,乙烯刺激后LRP上的elicitor-responsive protein 3-like isoform X1和参与糖酵解/糖异生代谢途径中的蛋白及酶类高积累,存在相似代谢调控机制的可能。糖酵解/糖异生代谢途径能够为酶促反应提供能量,并为橡胶链的延伸提供碳骨架[24],加速供能可能是LRP在乙烯促进天然橡胶生物合成过程中的作用之一。
乙烯促进了SRP上的内质网蛋白加工相关的蛋白积累,同时抑制了内吞过程相关的蛋白积累。单层膜包被的RP可能起源于内质网[25-26],SRP上参与橡胶生物合成的关键蛋白与内质网膜密切相关[27]。乙烯刺激可能通过促进SRP从内质网上招募更多的天然橡胶合成相关蛋白并进一步与SRP膜结合,进而增加橡胶产量。通过分析REF138和REF258的互作蛋白功能后发现,这一过程可能主要由REF138及其互作蛋白调控。有研究表明,乙烯刺激能够明显增加胶乳中SRP的比例[19],可能进一步促进天然橡胶的合成。
REF/SRPP蛋白家族成员可与其他参与天然橡胶合成调控的蛋白形成复合体结合在RP膜上。橡胶转移酶(HRT1,HRT2)、REF、SRPP及一种桥联蛋白(HRBP)以多种复合体形式结合在SRP上[28]。其中HRT1与HRBP形成的复合体具有异戊二烯基转移酶活性,并对脂膜及蛋白质α-螺旋结构域具有亲和力,介导蛋白与膜的结合过程[29]。本研究发现,REF138、REF175、REF258及SRPP117、SRPP204间存在相互作用的可能)。乙烯刺激后,LRP上积累量发生变化的REF/SRPP蛋白家族成员蛋白点有13个,SRPP上有22个,SRP上的蛋白复合体响应外源乙烯刺激后具备更为复杂和精细的天然橡胶合成调控机制。
针对RP膜上脂类化合物组成成分分析后发现,其磷脂含量远高于半乳糖苷含量[28]。本研究发现REF258与大量的脂代谢通路蛋白及相关酶类存在相互作用的可能,而乙烯刺激后LRP及SRP上的REF258不同亚型的积累均受到抑制。乙烯刺激可能通过改变RP膜的通透性影响跨膜运输,极有可能使某些相关蛋白与RP膜的结合位点发生改变,进而影响天然橡胶的合成调控。
外源乙烯的施用能够延长巴西橡胶树胶乳从乳管中流出的时间,进而增加单次割胶收集的橡胶产量[30]。胶乳凝集过程是影响胶乳流出时间的关键因素。巴西橡胶树胶乳LO中的胶乳凝集素样蛋白与SRPP结合,促进RP及LO的聚集进而导致胶乳凝集[31]。Hevein是胶乳中重要的凝固因子,贮存于LO中,一旦LO膜破碎释放后胶乳迅速凝集[32]。胶乳中的几丁质酶是一类抗凝固因子,能够提高胶乳稳定性延长排胶时间。乙烯刺激后,胶乳中几丁质酶活性随乙烯浓度增加而增加,胶乳产量较施用乙烯前也有所增加[33]。本研究发现REF138及REF258均可同胶乳中一种几丁质酶(hevamine-A)相互作用,乙烯刺激后,SRP上分子量为14 kDa左右的REF138蛋白亚型显著提升了积累水平,由于REF138同hevamine-A的相互作用,SRP上hevamine-A的积累水平可能也相应提升,缓解SRP间、SRP与LO的聚集,延缓胶乳凝集过程。
在本研究中,通过分离LRP及SRP并获得乙烯刺激后的蛋白样本进行差异分析,LRP上响应乙烯刺激发生积累差异的蛋白共计37个,SRP上共计56个。LRP的差异蛋白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,SRP的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。并通过Western Blot分析部分关键差异蛋白在胶乳不同组分间及相同组分乙烯刺激后的积累差异。对其中的REF138及REF258的互作蛋白进行检测,分别获得与REF138相互作用的25个以及与REF258存在互作关系的55个蛋白质信息,初步揭示了LRP和SRP响应外源乙烯刺激调控天然橡胶合成及增加胶乳产量的代谢调控机制。
  • 国家自然科学基金项目(31500557)
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2024年第45卷第6期
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doi: 10.3969/j.issn.1000-2561.2024.06.005
  • 接收时间:2023-08-01
  • 首发时间:2026-06-24
  • 出版时间:2024-06-25
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  • 收稿日期:2023-08-01
  • 修回日期:2023-09-08
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国家自然科学基金项目(31500557)
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    1.中国热带农业科学院热带生物技术研究所,海南海口 571101
    2.中国热带农业科学院海口实验站(热带果树研究所),海南海口 571101
    3.中国热带农业科学院橡胶研究所,海南儋州 571737

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* 仝征(TONG Zheng),E-mail:
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2种不同金属材料的力学参数

Family
属数
Number of
genus
种数
Number of
species
占总种数比例
Percentage of
total species (%)

Genus
种数
Number of
species
占总种数比例
Percentage of total
species (%)
鹅膏菌科Amanitaceae 2 11 5.26 鹅膏菌属 Amanita 10 4.78
小菇科 Mycenaceae 2 12 5.74 丝盖伞属 Inocybe 5 2.39
多孔菌科 Polyporaceae 8 14 6.70 蜡蘑属 Laccaria 5 2.39
红菇科 Russulaceae 3 23 11.00 小皮伞属 Marasmius 6 2.87
小菇属 Mycena 11 5.26
光柄菇属 Pluteus 5 2.39
红菇属 Russula 17 8.13
栓菌属 Trametes 5 2.39
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