Search results

根瘤农杆菌侵染植物过程中，至少有5种毒性蛋白（Vir）进入宿主细胞发挥作用，而其中VirD2与VirE2的作用最为关键，研究二者在水稻中的亚细胞定位，对农杆菌介导的水稻遗传转化机制的阐明具有重要意义。利用水稻叶鞘原生质体瞬时表达系统，发现3种冠瘿碱型的VirD2均只定位于细胞核中，与在拟南芥中相同；而3种冠瘿碱型的VirE2均主要定位于细胞核中，但在细胞质中仍有较多分布，与在拟南芥中的定位不同。因此，推测不同冠瘿碱型的农杆菌对水稻侵染能力的差异与VirD2和VirE2亚细胞定位的关系不大；同时表明根瘤农杆菌介导的拟南芥及水稻遗传转化机制存在相似性，但也有不同之处。 In the process of Agrobacterium tumefaciens-mediated infection of plants, at least 5 virulence (Vir) proteins play roles by entering the host cells, among which VirD2 and VirE2 are the key factors. Consequently, studying the sub-cellular localization of VirD2 and VirE2 in rice is crucial to understand the mechanism of Agrobacterium tumefaciens-mediated rice genetic transformation. Using the transient expression system of rice sheath protoplasts, we found that the VirD2 proteins of all 3 kinds of opine Agrobacterium are completely located in the nucleus, which is the same as in Arabidopsis; and the VirE2 proteins of all 3 kinds of opine Agrobacterium are mainly located in nucleus, but are also distributed in the cytoplasm, which

Summary The plant Artemisia annua is well known due to the production of artemisinin, a sesquiterpene lactone that is widely used in malaria treatment. Phytohormones play important roles in plant ... secondary metabolism, such as jasmonic acid ( JA ), which can induce artemisinin biosynthesis in A. annua . Nevertheless, the JA ‐inducing mechanism remains poorly understood. The expression of gene Aa MYC 2 was rapidly induced by JA and Aa MYC 2 binds the G‐box‐like motifs within the promoters of gene CYP 71 AV 1 and DBR 2 , which are key structural genes in the artemisinin biosynthetic pathway. Overexpression of Aa MYC 2 in A. annua significantly activated the transcript levels of CYP 71 AV 1 and DBR 2 , which resulted in an increased artemisinin content. By contrast, artemisinin content was reduced in the RNA i transgenic A. annua plants in which the expression of Aa MYC 2 was suppressed. Meanwhile, the RNA i transgenic A. annua plants showed lower sensitivity to methyl jasmonate treatment than the wild‐type plants. These results demonstrate that Aa MYC 2 is a positive regulator of artemisinin biosynthesis and is of great value in genetic engineering of A. annua for increased artemisinin production. more

Summary The glandular secretory trichomes ( GST s) on Artemisia annua leaves have the capacity to secrete and store artemisinin, a compound which is the most effective treatment for uncomplicated ... malaria. An effective strategy to improve artemisinin content is therefore to increase the density of GST s in A. annua . However, the formation mechanism of GST s remains poorly understood. To explore the mechanisms of GST initiation in A. annua , we screened myeloblastosis ( MYB ) transcription factor genes from a GST transcriptome database and identified a MIXTA transcription factor, Aa MIXTA 1, which is expressed predominantly in the basal cells of GST in A. annua . Overexpression and repression of Aa MIXTA 1 resulted in an increase and decrease, respectively, in the number of GST s as well as the artemisinin content in transgenic plants. Transcriptome analysis and cuticular lipid profiling showed that Aa MIXTA 1 is likely to be responsible for activating cuticle biosynthesis. In addition, dual‐luciferase reporter assays further demonstrated that Aa MIXTA 1 could directly activate the expression of genes related to cuticle biosynthesis. Taken together, Aa MIXTA 1 regulated cuticle biosynthesis and prompted GST initiation without any abnormal impact on the morphological structure of the GST s and so provides a new way to improve artemisinin content in this important medicinal plant. more

Artemisinin, a sesquiterpene lactone isolated from Artemisia annua L. (sweet wormwood), is extensively used in the treatment of malaria. In order to better understand the metabolism of terpenes in A. ... annua and the influence of terpene synthases on artemisinin yield, the expression pattern of a monoterpene alcohol dehydrogenase (ADH2) has been studied using transgenic plants expressing promoter–β‐glucuronidase (GUS) fusion. ADH2 played a major role in monoterpenoid biosynthesis including carveol, borneol, and artemisia ketone through in vitro biochemical analysis. In this study, the ADH2 promoter was cloned by the genome walking method. A number of putative cis‐acting elements were predicted in promoter region, suggesting that the ADH2 is driven by a complex regulation mechanism. ADH2 gene was highly expressed in old leaves, whereas the artemisinin biosynthetic genes were mainly expressed in bud and young leaves. The expression of ADH2 gene increased quickly during leaf development, revealed by qRT‐PCR. GUS expression analysis in different tissues of transgenic A. annua demonstrates that ADH2 expression is exclusively located to T‐shaped trichome, not glandular secretory trichome. more

Artemisinin is widely used as an antimalarial drug around the world. Artemisinic aldehyde Δ11(13) reductase (DBR2) is a key enzyme which reduces artemisinic aldehyde to dihydroartemisinic aldehyde in ... the biosynthesis of artemisinin. In this study, two fragments encompassing a putative promoter of DBR2, designated as DBR2pro1 and DBR2pro2, were isolated using genomic DNA walking. The transcription start site and the putative cis-elements of each version of promoter were predicted using bioinformatic analysis. In order to study the function of the cloned promoter, Artemisia annua was transformed with β-glucuronidase (GUS) reporter gene driven by DBR2pro1 and DBR2pro2, respectively. GUS staining results demonstrated that both DBR2pro1 and DBR2pro2 were strongly expressed in glandular secretory trichomes (GSTs) of leaf primordia and flower buds, but were not obviously expressed in roots, stems, old leaves, and fully developed flowers, thus indicating that the two versions of promoter were functional and specifically expressed in GSTs. more

Artemisinin, a sesquiterpene lactone isolated from Artemisia annua L. (sweet wormwood), is extensively used in the treatment of malaria. In order to better understand the metabolism of terpenes in A. ... annua and the influence of terpene synthases on artemisinin yield, the expression pattern of a monoterpene alcohol dehydrogenase (ADH2) has been studied using transgenic plants expressing promoter-beta-glucuronidase (GUS) fusion. ADH2 played a major role in monoterpenoid biosynthesis including carveol, borneol, and artemisia ketone through in vitro biochemical analysis. In this study, the ADH2 promoter was cloned by the genome walking method. A number of putative cis-acting elements were predicted in promoter region, suggesting that the ADH2 is driven by a complex regulation mechanism. ADH2 gene was highly expressed in old leaves, whereas the artemisinin biosynthetic genes were mainly expressed in bud and young leaves. The expression of ADH2 gene increased quickly during leaf development, revealed by qRT-PCR. GUS expression analysis in different tissues of transgenic A. annua demonstrates that ADH2 expression is exclusively located to T-shaped trichome, not glandular secretory trichome. more

Amorpha-4,11-diene synthase (ADS) is the first key enzyme of artemisinin biosynthetic pathway in Artemisia annua L. In this study, the promoter region of the ADS gene has been cloned and used to ... demonstrate the expression of GUS reporter gene in both glandular trichomes of A. annua and non-glandular trichomes of Arabidopsis thaliana following homologous and heterologous expression of ADS promoter–GUS fusion. Subsequently, 5′ sequential deletion analysis of the ADS promoter revealed that a short sequence, −350 upstream of the transcription start site, was sufficient for trichome-specific expression in A. thaliana and that the region from −350 to −300 contained essential elements for this observed specificity. However, frequencies of transgenic A. thaliana plants displaying trichome-specific expressions varied between different lines, and all the lines with deleted fragments of the ADS promoter showed lower frequencies than the line with full-length ADS promoter. Most lines with deleted ADS promoter–GUS fusions showed GUS expressions in the guard cells of stomata as well, which was not observed in A. thaliana plants transformed with the full-length ADS promoter. GUS activities varied among different transgenic lines as well, both in transiently transformed Nicotiana benthamiana and stably transformed A. thaliana, with promoter–deletion lines exhibiting higher GUS activities than the full-length ADS promoter line. more

Artemisinin is widely used as an antimalarial drug around the world. Artemisinic aldehyde Delta11(13) reductase (DBR2) is a key enzyme which reduces artemisinic aldehyde to dihydroartemisinic ... aldehyde in the biosynthesis of artemisinin. In this study, two fragments encompassing a putative promoter of DBR2, designated as DBR2pro1 and DBR2pro2, were isolated using genomic DNA walking. The transcription start site and the putative cis-elements of each version of promoter were predicted using bioinformatic analysis. In order to study the function of the cloned promoter, Artemisia annua was transformed with beta-glucuronidase (GUS) reporter gene driven by DBR2pro1 and DBR2pro2, respectively. GUS staining results demonstrated that both DBR2pro1 and DBR2pro2 were strongly expressed in glandular secretory trichomes (GSTs) of leaf primordia and flower buds, but were not obviously expressed in roots, stems, old leaves, and fully developed flowers, thus indicating that the two versions of promoter were functional and specifically expressed in GSTs. more