A de novo regulation design shows an effectiveness in altering plant secondary metabolism

Li, Mingzhuo; He, Xiping; Hovary, Christophe La; Zhu, Yue; Dong, Yilun; Liu, Shibiao; Xing, Hucheng; Liu, Yajun; Jie, Yucheng; Ma, Dongming; Yuzuak, Seyit; Xie, De‐Yu

doi:10.1016/j.jare.2021.06.017

Cited by 7 publications

(22 citation statements)

References 92 publications

(135 reference statements)

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“…AC elements were identified using the PLACE approach (https://www.dna.affrc.go.jp/PLACE/?action=newplace). Based on the sequences, promoters were cloned from A. annua as described before (Li et al 2022). Forward (5’ to 3’ end) and reverse (3’-5’) primer pairs (Supplementary Table S2) were designed to amplify at least 1 kb of nucleotides located immediately the upstream of the coding regions of the eight genes.…”

Section: Methodsmentioning

confidence: 99%

Molecular understanding of anthocyanin biosynthesis activated by PAP1 in engineered redArtemisia annuacells and regulation of 2, 4-dichlorophenoxyacetic acid

Dong

Cruz

et al. 2023

Preprint

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Artemisia annua is an effective antimalarial medicinal crop. We have established anthocyanin-producing red cell cultures from this plant with the overexpression of Production of Anthocyanin Pigment 1 (PAP1) encoding a R2R3MYB transcription factor. To understand the molecular mechanism by which PAP1 activated the entire anthocyanin pathway, we mined the genomic sequences of A. annua and obtained eight promoters of the anthocyanin pathway genes. Sequence analysis identified four types of AC cis-elements from six promoters, the MYB response elements (MRE) bound by PAP1. In addition, six promoters were determined to have at least one G-Box cis-element. Eight promoters were cloned for activity analysis. Duel luciferase assays showed that PAP1 significantly enhanced the promoting activity of seven promoters, indicating that PAP1 turned on the biosynthesis of anthocyanins via the activation of these pathway gene expression. To understand how 2,4-dichlorophenoxyacetic acid (2,4-D), an auxin, regulates the PAP1-activated anthocyanin biosynthesis, five different concentrations (0, 0.05, 0.5, 2.5, and 5 micro M) were tested to characterize anthocyanin production and profiles. The resulting data showed that the concentrations tested decreased the fresh weight of callus growth, anthocyanin levels, and the production of anthocyanins per petri dish. HPLC-qTOF-MS/MS based profiling showed that these concentrations did not alter anthocyanin profiles. Real time RT-PCR was completed to characterize the expression PAP1 and four representative pathway genes. The results showed that the five concentrations reduced the expression levels of the constitutive PAP1 transgene and three pathway genes significantly and eliminated chalcone synthase gene in expression either significantly or slightly. These data indicate that the constitutive PAP1 expression depends on gradients added in the medium. Based on these findings, the regulation of 2,4-D is discussed for anthocyanin engineering in red cells of A. annua.

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Section: Methodsmentioning

confidence: 99%

Molecular understanding of anthocyanin biosynthesis activated by PAP1 in engineered redArtemisia annuacells and regulation of 2, 4-dichlorophenoxyacetic acid

Dong

Cruz

et al. 2023

Preprint

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“…Anthocyanins are a group of pink/red/purple/blue flavonoid pigments with strong antioxidative activities that protect plants from stress-induced damages (Kano et al 2005; Yuzuak and Xie 2022; Shi and Xie 2014). To date, the biosynthesis of anthocyanins has gained intensive studies in biochemistry, genetics, metabolic engineering, and applications (Shi and Xie 2014; Holton and Cornish 1995; Li et al 2022; Zhang et al 2014). All pathway genes and most of regulatory genes have been characterized from both model and crop plants (Shi and Xie 2014; Khusnutdinov et al 2021).…”

Section: Introductionmentioning

confidence: 99%

“…In addition to pathway genes, most regulatory genes have been characterized from different plant species. Production of Anthocyanin Pigment 1 ( PAP1 ) that is one of master regulatory genes encodes a R2R3-MYB transcription factor (Borevitz et al 2000; Shi and Xie 2011; Li et al 2022). In Arabidopsis, it can enhance the expression of entry and early pathway genes and activate later pathway genes (Shi and Xie 2010; Shi and Xie 2011).…”

Section: Introductionmentioning

confidence: 99%

“…The AtPAP1 overexpression decreased alpha and beta acids in hops (Gatica-Arias et al 2012). AtPAP1 and Arabidopsis transparent testa 8 were introduced into commercial tobacco plants to decrease tobacco alkaloids and carcinogens (Li et al 2022). AtPAP1 was successfully used to engineer the anthocyanidin pathway of proanthocyanidins for animal nutrition (Xie and Dixon 2005).…”

Section: Introductionmentioning

confidence: 99%

“…Given that the formation of artemisinin requires the presence of oxidative condition rich in reactive oxygen species (ROS), the second goal was to understand whether A. annua cells rich in antioxidative anthocyanins could exclude the formation of artemisinin. We took advantage of AtPAP1, R2R3-MYB transformation factor (Xie et al 2006; Shi and Xie 2011; Li et al 2022), to engineer anthocyanins in A. annua cells. We developed four types of transgenic anthocyanin-producing A. annua (TAPA) cells, detected 17 anthocyanin or anthocyanidin compounds, from which four were identified to be cyanidin and pelargonidin, one cyanin, and one pelargonin, and characterized the biosynthesis of artemisinin in anthocyanin-producing TAPA cells.…”

Section: Introductionmentioning

confidence: 99%

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Metabolic engineering of anthocyanin pathway inArtemisia annuacells does not exclude the expression of artemisinin pathway

Judd

Dong

Sun

et al. 2023

Preprint

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Artemisia annua is the only medicinal crop to produce artemisinin for the treatment of malignant malaria. Unfortunately, hundreds of thousands of people still lose their life every year due to the lack of sufficient artemisinin. Artemisinin is considered to result from the spontaneous autoxidation of dihydroartemisinic acid in the presence of reactive oxygen species (ROS) in an oxidative condition of glandular trichomes (GTs),; however, whether increasing antioxidative compounds can inhibit artemisinin biosynthesis in plant cells is unknown. Anthocyanins are potent antioxidants that can remove ROS in plant cells. To date, no anthocyanins have been structurally elucidated from A. annua. In this study, our goals were to engineer anthocyanins in A. annua cells and to understand the artemisinin biosynthesis in anthocyanin-producing cells. Arabidopsis PAP1 (AtPAP1) was used to engineer four types of transgenic anthocyanin-producing A. annua (TAPA1 to 4) cells. Three wild type cell types were developed as controls. TAPA1 cells produced the highest contents of total anthocyanins. LC-MS analysis detected 17 anthocyanin or anthocyanidin compounds. Crystallization, LC/MS/MS and NMR analyses identified cyanidin, pelargonidin, one cyanin, and one pelargonin. An integrative analysis characterized that four types of TAPA cells expressed the artemisinin pathway and TAPA1 cells produced the highest artemisinin and artemisinic acid. The contents of arteannuin B were similar in seven cell types. These data showed that the engineering of anthocyanins does not eliminate the biosynthesis of artemisinin in cells. These data allow us to propose a new hypothesis that enzymes catalyze the formation of artemisinin from DHAA in non-GT cells. These findings show a new platform to increase artemisinin production via non-GT cells of A. annua.

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Metabolic engineering of the anthocyanin biosynthetic pathway in Artemisia annua and relation to the expression of the artemisinin biosynthetic pathway

Judd

Dong

Sun

et al. 2023

Planta

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A de novo regulation design shows an effectiveness in altering plant secondary metabolism

Cited by 7 publications

References 92 publications

Molecular understanding of anthocyanin biosynthesis activated by PAP1 in engineered redArtemisia annuacells and regulation of 2, 4-dichlorophenoxyacetic acid

Molecular understanding of anthocyanin biosynthesis activated by PAP1 in engineered redArtemisia annuacells and regulation of 2, 4-dichlorophenoxyacetic acid

Metabolic engineering of anthocyanin pathway inArtemisia annuacells does not exclude the expression of artemisinin pathway

Metabolic engineering of the anthocyanin biosynthetic pathway in Artemisia annua and relation to the expression of the artemisinin biosynthetic pathway

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