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

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.

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

There is a dichotomy in the biosynthetic pathway of terpenoid precursor isopentenyl diphosphate (IPP) in higher plant. One is the classical mevalonate pathway in cytosol, and the other is non-mevalonate pathway in plastid. To know the origin of the taxane ring system of taxol in suspension culture of Taxus chinensis, lovastatin and fosmidomycin were used to block the 3-hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) and 1-deoxy-D-xylulose-5-phosphate reducto-isomerase (DXR) in the mevalonate and non-mevalonate branch respectively of the terpenoid biosynthetic pathway. Methyl jasmonate (MJ) was used to improve the biosynthesis of taxol. Taxol content was determined by HPLC, the transcriptional expression of genes encoding DXR and HMGR were investigated by real time PCR. Taxol production was lowered by about 2/5 and 1/5 by fosmidomycin (200 mmol/L) and fosmidomycin (200 mmol/L)+MJ (100 mmol/L) treatment respectively, and was lowered by about 1/6 and 1/10 by lovastatin (1 mmol/L) and lovastatin (1 mmol/L) + MJ (100 mmol/L) respectively, which means that both mevalonate and non-mevalonate pathway contribute to taxol biosynthesis, and the latter is the main source of IPP. Inhibitors lovastatin and fosmidomycin both promoted the transcriptional expression of hmgr and dxr, which indicated a metabolic cross talk between cytosolic and plastidial pathways of taxol biosynthesis.