Yang Ke, Monfared Sajad Rashidi, Wang Hongzhen, Lundgren Anneli, Brodelius Peter E
Department of Chemistry and Biomedical Sciences, Linnaeus University, Kalmar, Sweden.
Plant Mol Biol. 2015 Jul;88(4-5):325-40. doi: 10.1007/s11103-015-0284-3. Epub 2015 Jan 24.
The artemisinic aldehyde double bond reductase (DBR2) plays an important role in the biosynthesis of the antimalarial artemisinin in Artemisia annua. Artemisinic aldehyde is reduced into dihydroartemisinic aldehyde by DBR2. Artemisinic aldehyde can also be oxidized by amorpha-4,11-diene 12-hydroxylase and/or aldehyde dehydrogenase 1 to artemisinic acid, a precursor of arteannuin B. In order to better understand the effects of DBR2 expression on the flow of artemisinic aldehyde into either artemisinin or arteannuin B, we determined the content of dihydroartemisinic aldehyde, artemisinin, artemisinic acid and arteannuin B content of A. annua varieties sorted into two chemotypes. The high artemisinin producers (HAPs), which includes the '2/39', 'Chongqing' and 'Anamed' varieties, produce more artemisinin than arteannuin B; the low artemisinin producers (LAPs), which include the 'Meise', 'Iran#8', 'Iran#14', 'Iran#24' and 'Iran#47' varieties, produce more arteannuin B than artemisinin. Quantitative PCR showed that the relative expression of DBR2 was significantly higher in the HAP varieties. We cloned and sequenced the promoter of the DBR2 gene from varieties of both the LAP and the HAP groups. There were deletions/insertions in the region just upstream of the ATG start codon in the LAP varities, which might be the reason for the different promoter activities of the HAP and LAP varieties. The relevance of promoter variation, DBR2 expression levels and artemisinin biosynthesis capabilities are discussed and a selection method for HAP varieties with a DNA marker is suggested. Furthermore, putative cis-acting regulatory elements differ between the HAP and LAP varieties.
青蒿醛双键还原酶（DBR2）在青蒿青蒿素生物合成中起重要作用。DBR2将青蒿醛还原为二氢青蒿醛。青蒿醛也可被紫穗槐-4,11-二烯12-羟化酶和/或醛脱氢酶1氧化为青蒿酸，即青蒿琥酯B的前体。为了更好地了解DBR2表达对青蒿醛流入青蒿素或青蒿琥酯B的影响，我们测定了分为两种化学型的青蒿品种中二氢青蒿醛、青蒿素、青蒿酸和青蒿琥酯B的含量。高青蒿素生产者（HAPs），包括“2/39”、“重庆”和“Anamed”品种，产生的青蒿素比青蒿琥酯B多；低青蒿素生产者（LAPs），包括“Meise”、“Iran#8”、“Iran#14”、“Iran#24”和“Iran#47”品种，产生的青蒿琥酯B比青蒿素多。定量PCR显示，DBR2在HAP品种中的相对表达显著更高。我们从LAP和HAP组的品种中克隆并测序了DBR2基因的启动子。LAP品种中ATG起始密码子上游区域存在缺失/插入，这可能是HAP和LAP品种启动子活性不同的原因。讨论了启动子变异、DBR2表达水平与青蒿素生物合成能力的相关性，并提出了一种用DNA标记选择HAP品种的方法。此外，HAP和LAP品种之间推定的顺式作用调控元件不同。
