磷甘霉素和洛伐它汀处理对中国红豆杉悬浮培养细胞生物合成紫杉醇的影响

Effects of Fosmidomycin and Lovastatin Treatment on Taxol Biosynthesis in Suspension Culture Cells of Taxus chinensis

采用非甲羟戊酸途径抑制剂磷甘霉素和甲羟戊酸途径抑制剂洛伐它汀对中国红豆杉悬浮细胞培养物进行处理。在添加和未添加茉莉酸甲酯诱导的情况下,前者使紫杉醇产量减少了2/5和1/5,后者使紫杉醇产量减少了1/6和1/10,表明两种途径对紫杉醇的生物合成都具有贡献,其中非甲羟戊酸途径贡献较大;通过定量PCR技术分别检测两条途径的关键酶5-磷酸脱氧木酮糖还原异构酶(DXR)和3-羟基-3-甲基戊二酰辅酶A还原酶(HMGR)mRNA水平的变化,发现两种抑制剂都能够激活hmgr和dxr的转录,表明两种代谢途径之间存在协同作用,共同为紫杉醇的生物合成提供前体。

There is a dichotomy in the biosyn-thetic pathway of terpenoid precursor isopentenyldiphosphate (IPP) in higher plant (Fig.1). One isthe classical mevalonate pathway in cytosol, andthe other is non-mevalonate pathway in plastid.To know the origin of the taxane ring system oftaxol in suspension culture of Taxus chinensis,lovastatin and fosmidomycin were used to blockthe 3-hydroxy-3-methylglutaryl-coenzyme A re-ductase (HMGR) and 1-deoxy-D-xylulose-5-phosphate reducto-isomerase (DXR) in themevalonate and non-mevalonate branch respec-tively of the terpenoid biosynthetic pathway.Methyl jasmonate (MJ) was used to improve thebiosynthesis of taxol. Taxol content was deter-mined by HPLC, the transcriptional expressionof genes encoding DXR and HMGR were in-vestigated by real time PCR. Taxol productionwas lowered by about 2/5 and 1/5 byfosmidomycin (200 μmol/L) and fosmidomycin (200 μmol/L)+MJ (100 μmol/L) treatmentrespectively, and was lowered by about 1/6 and1/10 by lovastatin (1 μmol/L) and lovastatin (1μmol/L) + MJ (100 μmol/L) respectively (Fig.2), which means that both mevalonate and non-mevalonate pathway contribute to taxolbiosynthesis, and the latter is the main source ofIPP. Inhibitors lovastatin and fosmidomycin bothpromoted the transcriptional expression of hmgrand dxr (Fig.4), which indicated a metaboliccross talk between cytosolic and plastidial path-ways of taxol biosynthesis.