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Isopentenyl Diphosphate Isomerase (IPI) Gene Silencing Negatively Affects Patchouli Alcohol Biosynthesis in Pogostemon cablin

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A Correction to this article was published on 13 November 2021

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Abstract

Pogostemon cablin is a medicinally and economically important plant because it contains a high content of patchouli oil. Patchouli oil is composed of more than 24 unique sesquiterpene compounds, with the most abundant being patchouli alcohol (PA). PA is a sesquiterpene and is mainly synthesized through the mevalonate (MVA) pathway in plants in which isopentenyl diphosphate isomerase (IPI) is a key enzyme. Here, a full-length cDNA encoding IPI (designated as PcIPI) was cloned from P. cablin through the rapid amplification of cDNA ends (RACE). Bioinformatics analysis revealed that the full-length PcIPI includes an 879 bp open reading frame (ORF) that encodes a 292 amino acid protein with a theoretical isoelectric point of 5.55 and a molecular mass of 32.8 kDa. Phylogenetic analysis demonstrated that PcIPI shared the most recent genetic relationship with the IPI of Isodon rubescens (86.43%), Salvia miltiorrhiza (85.38%), Plectranthus barbatus (86.54%), and Prunella vulgaris (86.31%). To verify the function of the PcIPI gene in PA biosynthesis, the P. cablin plants were infected with Agrobacterium tumefaciens bearing a pTRV2-PcIPI construct to induce virus-mediated PcIPI silencing. Quantitative real-time polymerase chain reaction (qRT-PCR) and gas chromatography mass spectrometry (GC-MS) analysis were applied to detect changes in the PcIPI catalytic products and transcription levels. The trend of decreasing transcript level was found to be similar to that of decreasing sesquiterpene content. These results revealed that PcIPI silencing had a negative effect on PA biosynthesis.

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Funding

This work was supported by grants from the National Natural Science Foundation of China (81660633 and 81860681) and the High level Talents Program of Hainan Natural Science Foundation (2019RC087).

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Authors and Affiliations

  1. College of Horticulture, Hainan University, Haikou, 570228, People’s Republic of China

    Wuping Yan, Yuzhang Yang, Yougen Wu, Jing Yu, Junfeng Zhang, Dongmei Yang & Zeeshan Ul Haq Muhammad

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  1. Wuping Yan
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  2. Yuzhang Yang
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  3. Yougen Wu
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  4. Jing Yu
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  5. Junfeng Zhang
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  6. Dongmei Yang
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  7. Zeeshan Ul Haq Muhammad
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Contributions

YGW conceived, designed, and supervised the research work. YZY and ZUHM performed the experiments. WPY and JY analyzed the data. WPY wrote the manuscript. JFZ and DMY provided inputs on data presentation and critically reviewed the manuscript. All the authors read and approved the final manuscript.

Corresponding author

Correspondence to Yougen Wu.

Additional information

Key Messages

• The isopentenyl diphosphate isomerase gene from Pogostemon cablin was cloned and characterized, and its function was explored using virus-induced gene silencing.

• Silencing of the PcIPI gene reduced the biosynthesis of patchouli alcohol.

• The IPI gene is a key gene in the sesquiterpene biosynthesis pathway, and tobacco rattle virus-mediated gene silencing can inhibit PcIPI gene expression.

Supplementary Information

Below is the link to the electronic supplementary material.

11105_2020_1269_MOESM1_ESM.tif

Supplementary file1 Figure S1. A. The vector genetic map of pTRV2. B. Schematic illustrations of the vector constructs used in this study. PcIPI fragment was inserted in pTRV2 vector. (TIF 109 KB)

11105_2020_1269_MOESM2_ESM.tif

Supplementary file2 Figure S2. The predicted conserved domains of PcIPI. A: Predicted by ORF of PcIPI; B: predicted by Coding sequence (CDS) of PcIPI. (TIF 1.54 MB)

Supplementary file3 Figure S3. Detection of pTRV1, pTRV2 and pTRV2-PcIPI fragments after VIGS infection. (TIF 103 KB)

Supplementary file4 Table S1. Primer sequences used in qRT-PCR. (DOCX 12 KB)

Supplementary file5 Table S2. Primers used to verify TRV transcription. (DOCX 12 KB)

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Yan, W., Yang, Y., Wu, Y. et al. Isopentenyl Diphosphate Isomerase (IPI) Gene Silencing Negatively Affects Patchouli Alcohol Biosynthesis in Pogostemon cablin. Plant Mol Biol Rep 39, 557–565 (2021). https://doi.org/10.1007/s11105-020-01269-0

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