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Molecular identification and expression of sesquiterpene pathway genes responsible for patchoulol biosynthesis and regulation in Pogostemon cablin.

ABSTRACT: BACKGROUND:Many commercially important drug and flavor compounds are secondary metabolites of terpenoid origin. Pogostemon cablin, a commercially important industrial and medicinal crop, accumulates abundant patchouli oil comprised of more than 24 unique sesquiterpene compounds, with the most abundant being patchouli alcohol. RESULTS:In this study, we analyzed the P. cablin transcriptome library, obtaining 74 terpenoid biosynthesis-related genes, and identified their expression patterns in leaves, stems, and flowers. These genes are members of 15 different families, and we detected all the enzymes involved in the sesquiterpenes pathway that are responsible for patchoulol biosynthesis. Sequence structure, homology, conserved domain properties, and phylogeny of certain identified genes were systematically investigated. Color complementation assay was used to verify the functional activity of the MEP pathway proteins. Exogenous hormone treatment revealed that patchoulol synthesis is induced by methyl jasmonate (MeJA). Quantitative reverse-transcription PCR analysis indicated that the MVA pathway genes (acetoacetyl-CoA thiolase, 3-hydroxy-3-methylglutaryl-coenzyme A reductase, mevalonate diphosphate decarboxylase, and farnesyl diphosphate synthase) participate in patchoulol biosynthesis and are mediated by MeJA. CONCLUSIONS:Taken together, this is the first report of integrated analysis of P. cablin MVA and MEP pathway related genes, providing a better understanding of terpenoid and/or patchoulol biosynthesis in P. cablin, and the basis for improving patchoulol production through genetic engineering.

SUBMITTER: Tang Y

PROVIDER: S-EPMC6606680 | biostudies-literature | 2019 Jul

REPOSITORIES: biostudies-literature

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Molecular identification and expression of sesquiterpene pathway genes responsible for patchoulol biosynthesis and regulation in Pogostemon cablin.

Tang Yun Y Zhong Liting L Wang Xiaobing X Zheng Hai H Chen Likai L

Botanical studies 20190702 1

<h4>Background</h4>Many commercially important drug and flavor compounds are secondary metabolites of terpenoid origin. Pogostemon cablin, a commercially important industrial and medicinal crop, accumulates abundant patchouli oil comprised of more than 24 unique sesquiterpene compounds, with the most abundant being patchouli alcohol.<h4>Results</h4>In this study, we analyzed the P. cablin transcriptome library, obtaining 74 terpenoid biosynthesis-related genes, and identified their expression pa ...[more]

PMID: 31267260

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Project description:BackgroundPogostemon cablin (Blanco) Benth. (Patchouli) is an important aromatic and medicinal plant and widely used in traditional Chinese medicine as well as in the perfume industry. Patchoulol is the primary bioactive component in P. cablin, its biosynthesis has attracted widespread interests. Previous studies have surveyed the putative genes involved in patchoulol biosynthesis using next-generation sequencing method; however, technical limitations generated by short-read sequencing restrict the yield of full-length genes. Additionally, little is known about the expression pattern of genes especially patchoulol biosynthesis related genes in response to methyl jasmonate (MeJA). Our understanding of patchoulol biosynthetic pathway still remained largely incomplete to date.ResultsIn this study, we analyzed the morphological character and volatile chemical compounds of P. cablin cv. 'Zhanxiang', and 39 volatile chemical components were detected in the patchouli leaf using GC-MS, most of which were sesquiterpenes. Furthermore, high-quality RNA isolated from leaves and stems of P. cablin were used to generate the first full-length transcriptome of P. cablin using PacBio isoform sequencing (Iso-Seq). In total, 9.7 Gb clean data and 82,335 full-length UniTransModels were captured. 102 transcripts were annotated as 16 encoding enzymes involved in patchouli alcohol biosynthesis. Accorded with the uptrend of patchoulol content, the vast majority of genes related to the patchoulol biosynthesis were up-regulated after MeJA treatment, indicating that MeJA led to an increasing synthesis of patchoulol through activating the expression level of genes involved in biosynthesis pathway of patchoulol. Moreover, expression pattern analysis also revealed that transcription factors participated in JA regulation of patchoulol biosynthesis were differentially expressed.ConclusionsThe current study comprehensively reported the morphological specificity, volatile chemical compositions and transcriptome characterization of the Chinese-cultivated P. cablin cv. 'Zhanxiang', these results contribute to our better understanding of the physiological and molecular features of patchouli, especially the molecular mechanism of biosynthesis of patchoulol. Our full-length transcriptome data also provides a valuable genetic resource for further studies in patchouli.

Project description:Pogostemon cablin, a medicinally and economically important perennial herb, is cultivated around the world due to its medicinal and aromatic properties. Different P. cablin cultivars exhibit different morphological traits and patchouli oil components and contents (especially patchouli alcohol (PA) and pogostone (PO)). According to the signature constituent of the leaf, P. cablin was classified into two different chemotypes, including PA-type and PO-type. To better understand the molecular mechanisms of PA biosynthesis, the transcriptomes of Chinese-cultivated P. cablin cv. PA-type "Nanxiang" (NX) and PO-type "Paixiang" (PX) were analyzed and compared with ribonucleic acid sequencing (RNA-Seq) technology. We obtained a total of 36.83 G clean bases from the two chemotypes, compared them with seven databases and revealed 45,394 annotated unigenes. Thirty-six candidate unigenes participating in the biosynthesis of PA were found in the P. cablin transcriptomes. Overall, 8,390 differentially expressed unigenes were identified between the chemotypes, including 2,467 upregulated and 5,923 downregulated unigenes. Furthermore, six and nine differentially expressed genes (DEGs) were mapped to the terpenoid backbone biosynthetic and sesquiterpenoid and triterpenoid biosynthetic pathways, respectively. One key sesquiterpene synthase gene involved in the sesquiterpenoid and triterpenoid biosynthetic pathways, encoding patchoulol synthase variant 1, was significantly upregulated in NX. Additionally, GC-MS analysis of the two chemotypes in this study showed that the content of PA in NX was significantly higher than that of PX, while the content of PO showed the opposite phenotype. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that the DEG expression tendency was consistent with the transcriptome sequencing results. Overall, 23 AP2/ERF, 13 bHLH, 11 MYB, 11 NAC, three Trihelix, 10 WRKY and three bZIP genes that were differentially expressed may act as regulators of terpenoid biosynthesis. Altogether, 8,314 SSRs were recognized within 6,825 unigenes, with a distribution frequency of 18.32%, among which 1,202 unigenes contained more than one SSR. The transcriptomic characteristics of the two P. cablin chemotypes are comprehensively reported in this study, and these results will contribute to a better understanding of the molecular mechanism of PA biosynthesis. Our transcriptome data also provide a valuable genetic resource for further studies on P. cablin.

Dataset Information

Molecular identification and expression of sesquiterpene pathway genes responsible for patchoulol biosynthesis and regulation in Pogostemon cablin.

Publications

Molecular identification and expression of sesquiterpene pathway genes responsible for patchoulol biosynthesis and regulation in Pogostemon cablin.

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