通过胶乳转录组分析探析巴西橡胶树橡胶生物合成的研究

Insights into Rubber Biosynthesis from Transcriptome Analysis of Hevea Brasiliensis Latex

巴西橡胶树(Hevea brasiliensis)是天然橡胶(顺式-聚异戊二烯)商业化生产中栽培最为广泛的一个物种。本研究中,为了深入分析并了解胶乳转录谱,尤其是橡胶生物合成相关基因的转录表达情况,我们从橡胶树的胶乳这种代表单一细胞类型的胞质成分中分离得到10 040个表达序列标签(ESTs),经ESTs序列编辑组装之后共获得3 441个独立转录本(UTs)。基于标准基因词汇体系对这些UTs进行功能分类的结果显示,它们中的73.8%均与未知功能的基因相关。在高度表达的ESTs中,编码橡胶生物合成及应激或防御相关蛋白的ESTs占有相当大的比例,其中编码分属三个蛋白质家族的橡胶粒子膜蛋(RPMPs)的序列其所占ESTs的比例即高达12%。从这些ESTs中还鉴定出了九种RPMP异型体(7.9～27KDa),其中包括14 Kda的REF(橡胶延伸因子)及22 Kda的SRPP(小橡胶粒子蛋白)。试验中利用抗REF及SRPP抗体也检测到了胶乳中多种RPMP异型体的表达。此外,EST及定量逆转录-多聚酶链式反(QRT-PCR)分析均证实REF和SRPP为胶乳中最丰富的转录产物。除橡胶生物合成之外,比较序列分析结果显示,RPMPs与植物界存在的应激相关蛋白序列具有高度相似性。最后,本文还根据转录本丰度及差异基因表达就RPMP在顺式-聚异戊二烯生物合成中的功能作了进一步的探讨和推测。

Hevea brasiliensis is the most widely cultivated species for commercial production of natural rubber（cis—polyisoprene）.In this study,-10 040 expressed sequence tags（ESTs） were generated from the latex of the rubber tree,which represents the cytoplasmic content of a single cell type,in order to analyse the latex transcription profile with emphasis on rubber biosynthesis—related genes.A total of 3 441 unique transcripts（UTs） were obtained after quality editing and assembly of EST sequences. Functional classification of UTs according to the Gene Ontology convention showed that 73. 8%were related to genes of unknown function.Among highly expressed ESTs,a significant proportion encoded proteins related to rubber biosynthesis and stress or defense responses.Sequences encoding rubber particle membrane proteins（RPMPs） belonging to three protein families accounted for 12%of the ESTs.Characterization of these ESTs revealed nine RPMP variants（7.9—27 kDa） including the 14 kDa REF（rubber elongation factor） and 22 kDa SRPP（small rubber particle protein）. The expression of multiple RPMP isoforms in latex was shown using antibodies against REF and SRPP. Both EST and quantitative reverse transcription—PCR（QRTPCR） analyses demonstrated REF and SRPP to be the most abundant transcripts in latex.Besides rubber biosynthesis,comparative sequence analysis showed that the RPMPs are highly similar to sequences in the plant kingdom having stress—related functions.Implications of the RPMP function in cis—polyisoprene biosynthesis in the context of transcript abundance and differential gene expression are discussed.