Division of Forest Resources, Kangwon National University, Chunchon 200-701, Republic of Korea.
Plant Cell Physiol. 2011 Jan;52(1):125-37. doi: 10.1093/pcp/pcq179. Epub 2010 Dec 5.
Squalene synthase (SQS) catalyzes the biosynthesis of squalene by condensing two molecules of farnesyl pyrophosphate (FPP), a key precursor in sterol and triterpene biosynthesis. Previously, we reported that PgSS1 overexpression results in the enhanced biosynthesis of both phytosterols and triterpene saponins in Panax ginseng. Here, cDNAs encoding two new SQS homologs (PgSS2 and PgSS3) from a P. ginseng expressed sequence tag (EST) library are described. Functional complementation analysis revealed that ectopic expression of PgSS1, PgSS2 and PgSS3 in the yeast erg9 mutant strain 2C1 lacking SQS activity restored ergosterol prototrophy. The recombinant mutant yeast produced squalene, squalene epoxide and ergosterol. PgSS1 (mRNA) was highly transcribed in all organs, whereas PgSS2 and PgSS3 (mRNAs) were only transcribed in specific organs. All three genes were activated positively by an elicitor (methyl jasmonate), but their transcriptional patterns were different. In situ hybridization analysis revealed that both PgSS1 and PgSS3 transcripts were preferentially accumulated near conducting tissue in the petiole. The PgSS1 and PgSS3 promoters were isolated, and the tissue- and organ-specific regulation of PgSS genes was examined. Transgenic ginseng was constructed by introducing PgSS1 and PgSS3 promoters fused to the β-glucuronidase (GUS) gene. GUS expression driven by the PgSS1 promoter was found in both roots and shoots, but PgSS3-driven GUS was only found in shoots. These results suggest that the three SQS genes are differently expressed and that all three SQS enzymes are involved in squalene production in P. ginseng.
鲨烯合酶（SQS）通过缩合两个法呢基焦磷酸（FPP）分子催化鲨烯的生物合成，FPP 是甾醇和三萜生物合成的关键前体。以前，我们报道过 PgSS1 的过表达导致人参中植物甾醇和三萜皂苷的生物合成增强。在这里，我们描述了从人参表达序列标签（EST）文库中编码两个新的 SQS 同源物（PgSS2 和 PgSS3）的 cDNA。功能互补分析表明，在缺乏 SQS 活性的酵母 erg9 突变体 2C1 中异位表达 PgSS1、PgSS2 和 PgSS3 恢复了麦角固醇原养型。重组突变酵母产生了鲨烯、鲨烯环氧化物和麦角固醇。PgSS1（mRNA）在所有器官中均高度转录，而 PgSS2 和 PgSS3（mRNA）仅在特定器官中转录。所有三种基因均被诱导剂（茉莉酸甲酯）正向激活，但它们的转录模式不同。原位杂交分析表明，PgSS1 和 PgSS3 转录本在叶柄的导组织附近优先积累。分离了 PgSS1 和 PgSS3 启动子，并检查了 PgSS 基因的组织和器官特异性调节。通过引入 PgSS1 和 PgSS3 启动子与β-葡萄糖醛酸酶（GUS）基因融合构建了转基因人参。发现 PgSS1 启动子驱动的 GUS 表达存在于根和地上部分，但 PgSS3 驱动的 GUS 仅存在于地上部分。这些结果表明，这三个 SQS 基因表达不同，所有三种 SQS 酶都参与人参中鲨烯的产生。
