Molecular cloning and expression analysis of a squalene synthase gene from a medicinal plant, Euphorbia pekinensis Rupr.

Zheng, Zhujun; Cao, Xiaoying; Li, Changgen; Chen, Yongqiang; Yuan, Bo; Xu, Yan; Jiang, Jihong

doi:10.1007/s11738-013-1333-7

Cited by 17 publications

(5 citation statements)

References 29 publications

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“…Some studies found that the tissue expression pattern of SQS varies greatly in different plants. Similar patterns of gene expression were found in S. miltiorrhiza [ 22 ], Euphorbia pekinensis [ 36 ], Taxus cuspidata [ 32 ], while some different expression patterns were found in B. platyphylla [ 37 ], W. somnifera [ 38 ], and Euphorbia tirucalli [ 39 ]. In addition, several studies have reported that the MeJA could upregulates the expression of SQS [ 21 , 36 ].…”

Section: Discussionmentioning

confidence: 64%

Cloning, Expression Analysis and Functional Characterization of Squalene Synthase (SQS) from Tripterygium wilfordii

et al. 2018

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Celastrol is an active triterpenoid compound derived from Tripterygium wilfordii which is well-known as a traditional Chinese medicinal plant. Squalene synthase has a vital role in condensing two molecules of farnesyl diphosphate to form squalene, a key precursor of triterpenoid biosynthesis. In the present study, T. wilfordii squalene synthase (TwSQS) was cloned followed by prokaryotic expression and functional verification. The open reading frame cDNA of TwSQS was 1242 bp encoding 413 amino acids. Bioinformatic and phylogenetic analysis showed that TwSQS had high homology with other plant SQSs. To obtain soluble protein, the truncated TwSQS without the last 28 amino acids of the carboxy terminus was inductively expressed in Escherichia coli Transetta (DE3). The purified protein was detected by SDS-PAGE and Western blot analysis. Squalene was detected in the product of in vitro reactions by gas chromatograph-mass spectrometry, which meant that TwSQS did have catalytic activity. Organ-specific and inducible expression levels of TwSQS were detected by quantitative real-time PCR. The results indicated that TwSQS was highly expressed in roots, followed by the stems and leaves, and was significantly up-regulated upon MeJA treatment. The identification of TwSQS is important for further studies of celastrol biosynthesis in T. wilfordii.

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Section: Discussionmentioning

confidence: 64%

Cloning, Expression Analysis and Functional Characterization of Squalene Synthase (SQS) from Tripterygium wilfordii

et al. 2018

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“…In plants, like many other enzymes of the MVA pathway, is encoded by small gene families. A single SQS gene has been reported in many plants, such as Taxus cuspidata ( Huang et al, 2007 ), Euphorbia tirucalli ( Uchida et al, 2009 ), Oryza sativa ( Hata et al, 1997 ), Lotus japonicus ( Akamine et al, 2003 ), Chlorophytum borivilianum ( Kalra et al, 2013 ), and E. pekinensis ( Zheng et al, 2013 ), while it has been found that there are two SQS genes in Arabidopsis thaliana ( Kribii et al, 1997 ), tobacco ( Devarenne et al, 1998 ), Glycyrrhiza glabra ( Hayashi et al, 1999 ), Withania somnifera ( Gupta et al, 2012 ), Glycine max ( Nguyen et al, 2013 ), and Salvia miltiorrhiza Bunge ( Ma et al, 2014 ), and three SQS genes in Panax ginseng ( Kim et al, 2011 ).…”

Section: Introductionmentioning

confidence: 99%

Molecular Cloning and Functional Analysis of Squalene Synthase 2(SQS2) in Salvia miltiorrhiza Bunge

et al. 2016

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Salvia miltiorrhiza Bunge, which is also known as a traditional Chinese herbal medicine, is widely studied for its ability to accumulate the diterpene quinone Tanshinones. In addition to producing a variety of diterpene quinone, S. miltiorrhiza Bunge also accumulates sterol, brassinosteroid and triterpenoids. During their biosynthesis, squalene synthase (SQS, EC 2.5.1.21) converts two molecules of the hydrophilic substrate farnesyl diphosphate (FPP) into a hydrophobic product, squalene. In the present study, cloning and characterization of S. miltiorrhiza Bunge squalene synthase 2 (SmSQS2, Genbank Accession Number: KM408605) cDNA was investigated subsequently followed by its recombinant expression and preliminary enzyme activity. The full-length cDNA of SmSQS2 was 1 597 bp in length, with an open reading frame of 1 245 bp encoding 414 amino acids. The deduced amino acid sequence of SmSQS2 shared high similarity with those of SQSs from other plants. To obtain soluble recombinant enzymes, the truncated SmSQS2 in which 28 amino acids were deleted from the carboxy terminus was expressed as GST-Tag fusion protein in Escherichia coli BL21 (DE3) and confirmed by SDS-PAGE and Western Blot analysis, and the resultant bacterial crude extract was incubated with FPP and NADPH. Gas chromatograph-mass spectrometer analysis showed that squalene was detected in the in vitro reaction mixture. The gene expression level was analyzed through Quantitative real-time PCR, and was found to be higher in roots as compared to the leaves, and was up-regulated upon YE+ Ag+ treatment. These results could serve as an important to understand the function of the SQS family. In addition, the identification of SmSQS2 is important for further studies of terpenoid and sterol biosynthesis in S. miltiorrhiza Bunge.

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“…Squalene is biosynthesized by combining two molecules of FPP by squalene synthase. A squalene synthase gene was isolated from Euphorbia pekinensis [ 62 ] and Euphorbia tirucalli [ 63 ]. We found two copies of squalene synthase in E. maculata , and both copies were actively expressed in the three organs.…”

Section: Discussionmentioning

confidence: 99%

Identifying Terpenoid Biosynthesis Genes in Euphorbia maculata via Full-Length cDNA Sequencing

Jeon

Roy²,

Choi³

et al. 2022

Molecules

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The annual herb Euphorbia maculata L. produces anti-inflammatory and biologically active substances such as triterpenoids, tannins, and polyphenols, and it is used in traditional Chinese medicine. Of these bioactive compounds, terpenoids, also called isoprenoids, are major secondary metabolites in E. maculata. Full-length cDNA sequencing was carried out to characterize the transcripts of terpenoid biosynthesis reference genes and determine the copy numbers of their isoforms using PacBio SMRT sequencing technology. The Illumina short-read sequencing platform was also employed to identify differentially expressed genes (DEGs) in the secondary metabolite pathways from leaves, roots, and stems. PacBio generated 62 million polymerase reads, resulting in 81,433 high-quality reads. From these high-quality reads, we reconstructed a genome of 20,722 genes, in which 20,246 genes (97.8%) did not have paralogs. About 33% of the identified genes had two or more isoforms. DEG analysis revealed that the expression level differed among gene paralogs in the leaf, stem, and root. Whole sets of paralogs and isoforms were identified in the mevalonic acid (MVA), methylerythritol phosphate (MEP), and terpenoid biosynthesis pathways in the E. maculata L. The nucleotide information will be useful for identifying orthologous genes in other terpenoid-producing medicinal plants.

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Molecular cloning and expression analysis of a squalene synthase gene from a medicinal plant, Euphorbia pekinensis Rupr.

Cited by 17 publications

References 29 publications

Cloning, Expression Analysis and Functional Characterization of Squalene Synthase (SQS) from Tripterygium wilfordii

Cloning, Expression Analysis and Functional Characterization of Squalene Synthase (SQS) from Tripterygium wilfordii

Molecular Cloning and Functional Analysis of Squalene Synthase 2(SQS2) in Salvia miltiorrhiza Bunge

Identifying Terpenoid Biosynthesis Genes in Euphorbia maculata via Full-Length cDNA Sequencing

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