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Molecular cloning, characterization and expression analysis of two 12-oxophytodienoate reductases (NtOPR1 and NtOPR2) from Nicotiana tabacum

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Abstract

Background

12-oxophytodienoic acid (OPDA) is a signaling molecule involved in defense and stress responses in plants. 12-oxophytodienoate reductase (OPR) is involved in the biosynthesis of jasmonic acid and trigger the conversion of OPDA into 3-oxo-2(2′[Z]-pentenyl)-cyclopentane-1-octanoic acid (OPC-8:0).

Methods and results

Sequence analysis revealed that Nicotiana tabacum 12-oxophytodienoate reductase 1 (OPR1) and OPR2 encoded polypeptides of 375 and 349 amino acids with molecular masses of 41.67 and 39.04 kilodaltons (kDa), respectively, while the deduced protein sequences of NtOPR1 and NtOPR2 showed high homology with other 12-oxophytodienoate reductases. BLAST (Basic local alignment search tool) analysis revealed that both NtOPRs belong to the family of Old Yellow Enzymes (OYE), and analysis of genomic DNA structure indicated that both genes include 5 exons and 4 introns. Phylogenetic analysis using MEGA X showed that NtOPR1 and NtOPR2 shared a close evolutionary relationship with Nicotiana attenuata 12-oxophytodienoate reductases. In silico analysis of subcellular localization indicated the probable locations of NtOPR1 and NtOPR2 to be the cytoplasm and the peroxisome, respectively. Tissue-specific expression assays via qRT-PCR revealed that NtOPR1 and NtOPR2 genes were highly expressed in Nicotiana tabacum roots, temperately expressed in leaves and flowers, while low expression was observed in stem tissue.

Conclusions

Presently, two 12-oxophytodienoate reductase genes (NtOPR1 and NtOPR2) were cloned and comprehensively characterized. Our findings provide comprehensive analyses that may guide future deep molecular studies of 12-oxophytodienoate reductases in Nicotiana tabacum.

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Funding

This work was supported by the Science and Technology Plan Project of Yunnan Provincial Tobacco Company (2018530000241016, 2018530000241020, 2019530000241028, 2019530000241011), Shanghai Tobacco Group Co., Ltd (20193100001, 20203100001), Yunnan Provincial Department of Science and Technology (2019FH001(-051)), and Science and Technology Project of Yunnan Provincial Education Department (2021J0716).

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Authors and Affiliations

  1. College of Agriculture and Life Sciences, Yunnan Urban Agricultural Engineering & Technological Research Center, Kunming University, Kunming, China

    Feiyan Huang, Lei Yu & Yanguo Ke

  2. The Research Center for Ornamental Plants, College of Forestry and Landscape Architecture, South China Agricultural University, Guangzhou, 510642, China

    Farhat Abbas

  3. College of Horticulture Science, South China Agricultural University, Guangzhou, 510642, China

    Dylan O’Neill Rothenberg

  4. Department of Crop Science and Technology, College of Agriculture, South China Agricultural University, Guangzhou, 510642, China

    Muhammad Imran

  5. Department of Plant Breeding and Genetics, University of Haripur, Khyber Pakhtunkhwa, Pakistan

    Sajid Fiaz

  6. State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, Guangdong Provincial Key Laboratory of Plant Molecular Breeding, South China Agricultural University, Guangzhou, 510642, China

    Naveed ur Rehman

  7. College of Horticulture and Landscape Architecture, Northeast Agricultural University, Harbin, 150030, Heilongjiang, China

    Sikandar Amanullah

  8. Department of Botany, Lahore College for Women University, Lahore, Pakistan

    Afifa Younas

  9. Material Procurement Center, Shanghai Tobacco Group Co., Ltd, Shanghai, 200082, Yunnan, China

    Yan Ding & Xianjie Cai

  10. Tobacco Leaf Purchase Center, China Tobacco Henan Industrial Co., Ltd., Zhengzhou, 450000, China

    Xiaolong Chen

  11. Kunming Tobacco Corporation of Yunnan Province, Kunming, 650021, China

    Xianwen Ye

  12. Honghe Tobacco Corporation of Yunnan Province, Honghe, 661400, China

    Lin Jiang

  13. College of Economics and Management, Kunming University, Kunming, China

    Yanguo Ke

  14. Lincang Company of Yunnan Tobacco Company, Lincang, 677000, China

    Yuansheng He

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Huang, F., Abbas, F., Rothenberg, D.O. et al. Molecular cloning, characterization and expression analysis of two 12-oxophytodienoate reductases (NtOPR1 and NtOPR2) from Nicotiana tabacum. Mol Biol Rep 49, 5379–5387 (2022). https://doi.org/10.1007/s11033-022-07114-9

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