Advertisement

Isolation and characterization of a 3-hydroxy-3-methylglutaryl coenzyme A reductase 2 promoter from Salvia miltiorrhiza

  • Piotr Szymczyk
  • Renata Grąbkowska
  • Ewa Skała
  • Marta Żebrowska
  • Ewa Balcerczak
  • Agnieszka Jeleń
Original Article
  • 154 Downloads

Abstract

The aim of the present study was to isolate and characterize the 5′ regulatory region of Salvia miltiorrhiza 3-hydroxy-3-methylglutaryl coenzyme A reductase 2 gene. The entire fragment is 2696 bp long and consists of the promoter, 5′UTR and 85 nucleotide 5′ fragments of the CDS region. The results of in silico bioinformatic tests indicate that the promoter region contains repetitions of many potential cis-active elements serving as the recognition sites for transcription factors. Data obtained from the in silico tests are verified by co-expression studies based on A. thaliana microarray data to make them more probably to occur. The bioinformatic analysis indicated no tandem repeats or CpNpG islands in the promoter. However, potential target sites for miRNA 156 and miRNA 159 were found in the 5′ UTR segment. In addition, two possible polymorphic sites, A2719G and A2744C, were found in the CDS region. Finally, the activity of isolated fragment was evaluated experimentally by quantitative RT–PCR. The promoter activity of the isolated 2696 bp HMGR2 gene fragment was confirmed by RT–PCR studies of in vitro cultured, transformed S. miltiorrhiza plants. Analysis of the RT–PCR results revealed temporal changes in the promoter activity occurring in response to treatment by five abiotic factors: auxin, gibberellin, salicylic acid, abscisic acid and 100 mM NaCl.

Keywords

miRNA Polymorphic site Promoter RT–PCR Trans-factor 

Abbreviations

ABA

Abscisic acid

CDS

Coding DNA sequence

IAA

Indole-3-aceticacid

GA

Gibberellic acid

HMGR

3-hydroxy-3-methylglutaryl coenzyme A reductase

IAA

Indole-3-aceticacid

MeJ

Methyl jasmonate

PP2A

Ser/Thr protein phosphatase 2A

5′UTR

5′ untranslated region

Notes

Acknowledgements

Authors are extremely grateful to Dean Elżbieta Mikiciuk-Olasik, the former Dean of the Faculty of Pharmacy, Medical University of Łódź, and Dean Daria Orszulak-Michalak for providing financial support. Authors gratefully acknowledge the technical assistance of Wacław Prószyński.

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflict of interest.

Supplementary material

13562_2017_434_MOESM1_ESM.docx (2.2 mb)
Supplementary material 1 (DOCX 2207 kb)

References

  1. Akhtar N, Gupta P, Sangwan NS, Sangwan RS, Trivedi PK (2013) Cloning and functional characterization of 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Withania somnifera: an important medicinal plant. Protoplasma 250:613–622CrossRefPubMedGoogle Scholar
  2. Berg RH (2004) Evaluation of spectral imaging for plant cell analysis. J Microsc 214:174–181CrossRefPubMedGoogle Scholar
  3. Birnboim HC, Doly J (1979) A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res 7:1513–1523CrossRefPubMedPubMedCentralGoogle Scholar
  4. Bolle C, Sopory S, Lubberstedt T, Herrmann RG, Oelmuller R (1994) Segments encoding 5′-untranslated leaders of genes for thylakoid proteins contain cis-elements essential for transcription. Plant J 6:513–523CrossRefPubMedGoogle Scholar
  5. Cao X, Jacobsen SE (2002) Locus-specific control of asymmetric and CpNpG methylation by the DRM and CMT3 methyltransferase genes. Proc Natl Acad Sci USA 4:16491–16498CrossRefGoogle Scholar
  6. Cao XY, Li Ch-G, Mao Q, Zheng ZJ, Jiang JH (2011) Molecular cloning and expression analysis of a leaf-specific expressing 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase gene from Michelia chapensis Dandy. J Med Plant Res 5:3868–3875Google Scholar
  7. Chow ChN, Zheng HQ, Wu NY, Chien ChH, Huang HD, Lee TY et al (2016) PlantPAN 2.0: an update of plant promoter analysis navigator for reconstructing transcriptional regulatory networks in plants. Nucleic Acids Res 44:D1154–D1160CrossRefPubMedGoogle Scholar
  8. Ckurshumova W, Caragea AE, Goldstein RS, Berleth T (2011) Glow in the dark: fluorescent proteins as cell and tissue-specific markers in plants. Mol Plant 4:794–804CrossRefPubMedGoogle Scholar
  9. Cormack BP, Valdivia RH, Falkow S (1996) FACS-optimized mutants of the green fluorescent protein (GFP). Gene 173:33–38CrossRefPubMedGoogle Scholar
  10. Dai Z, Cui G, Zhou S-F, Zhang X, Huang L (2011) Cloning and characterization of a novel 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Salvia miltiorrhiza involved in diterpenoid tanshinone accumulation. J Plant Physiol 168:148–157CrossRefPubMedGoogle Scholar
  11. Daraselia ND, Svetlana Tarchevskaya S, Narita JO (1996) The promoter for tomato 3-hydroxy-3-methylglutaryl coenzyme a reductase gene 2 has unusual regulatory elements that direct high-leve1 expression. Plant Physiol 112:727–733CrossRefPubMedPubMedCentralGoogle Scholar
  12. Friesen JA, Rodwell VW (2004) The 3-hydroxy-3-methylglutaryl coenzyme-A (HMG-CoA) reductases. Genome Biol 5:e248CrossRefGoogle Scholar
  13. Griffiths-Jones S, Grocock RJ, van Dongen S, Bateman A, Enright AJ (2006) miRbase: microRNA sequences, targets and gene nomenclature. Nucleic Acids Res 34:D140–D144CrossRefPubMedGoogle Scholar
  14. Guo D, Zhou Y, Li HL, Zhu JH, Wang Y, Chen XT, Peng SQ (2017) Identification and characterization of the abscisic acid (ABA) receptor gene family and its expression in response to hormones in the rubber tree. Sci Rep-UK 7:45157CrossRefGoogle Scholar
  15. Ha SH, Kim JB, Hwang YS, Lee SW (2003) Molecular characterization of three 3-hydroxy-3-methylglutaryl-CoA reductase genes including pathogen-induced Hmg2 from pepper (Capsicum annuum). Biochim Biophys Acta 1625:253–260CrossRefPubMedGoogle Scholar
  16. Hellens R, Mullineaux P, Klee H (2000) Technical Focus: a guide to Agrobacterium binary Ti vectors. Trends Plant Sci 5:446–451CrossRefPubMedGoogle Scholar
  17. Jain AK, Vincent RM, Nessler CL (2000) Molecular characterization of a hydroxymethylglutaryl-CoA reductase gene from mulberry (Morus alba L.). Plant Mol Biol 42:559–569CrossRefPubMedGoogle Scholar
  18. Jing F, Zhang L, Li M, Tang Y, Wang Y, Wang Y, Wang Q, Pan Q, Wang G, Tang K (2009) Abscisic acid (ABA) treatment increases artemisinin content in Artemisia annua by enhancing the expression of genes in artemisinin biosynthetic pathway. Biologia 64:319–323CrossRefGoogle Scholar
  19. Kai G, Liao P, Zhou W, Wang J, Xu H, Liu Y, Zhang L (2010) Characterization, expression profiling, and functional identification of a gene encoding geranylgeranyl diphosphate synthase from Salvia miltiorrhiza. Biotechnol Bioprocess Eng 15:236–245CrossRefGoogle Scholar
  20. Kang MK, Park KS, Choi D (1998) Coordinated expression of defense-related genes by TMV infection or salicylic acid treatment in tobacco. Mol Cells 8:388–392PubMedGoogle Scholar
  21. Karimi M, Inzé D, Depicker A (2002) GATEWAY vectors for Agrobacterium-mediated plant transformation. Trends Plant Sci 7:193–195CrossRefPubMedGoogle Scholar
  22. Kawoosa T, Gahlan P, Devi AS, Kumar S (2014) The GATA and SORLIP motifs in the 3-hydroxy-3-methylglutaryl-CoA reductase promoter of Picrorhiza kurrooa for the control of light-mediated expression. Funct Integr Genom 14:191–203CrossRefGoogle Scholar
  23. Khan S, Qureshi MI, Kamaluddin Alam T, Abdin MZ (2007) Protocol for isolation of genomic DNA from dry and fresh roots of medicinal plants suitable for RAPD and restriction digestion. Afr J Biotechnol 6:175–178Google Scholar
  24. Kibbe WA (2007) OligoCalc: an online oligonucleotide properties calculator. Nucleic Acids Res 35W:W43–W46CrossRefGoogle Scholar
  25. Kirby J, Keasling JD (2009) Biosynthesis of plant isoprenoids: perspectives for microbial engineering. Ann Rev Plant Biol 60:335–355CrossRefGoogle Scholar
  26. Law JA, Jacobsen SE (2010) Establishing, maintaining and modifying DNA methylation patterns in plants and animals. Nat Rev Genet 11:204–220CrossRefPubMedPubMedCentralGoogle Scholar
  27. Leivar P, Antolín-Llovera M, Ferrero S, Closa M, Arró M, Ferrer A, Boronat A, Campos N (2011) Multilevel control of Arabidopsis 3-hydroxy-3-methylglutaryl coenzyme A reductase by protein phosphatase 2A. Plant Cell 23:1494–1511CrossRefPubMedPubMedCentralGoogle Scholar
  28. Li H, Ruan J, Durbin R (2008) Mapping short DNA sequencing reads and calling variants using mapping quality scores. Genome Res 18:1851–1858CrossRefPubMedPubMedCentralGoogle Scholar
  29. Li SB, Xie ZZ, Hu ChG, Zhang JZ (2016) A review of auxin response factors (ARFs) in plants. Front Plant Sci 7:47PubMedPubMedCentralGoogle Scholar
  30. Liao P, Zhou W, Zhang L, Wang J, Yan X, Zhang Y, Zhang R, Li L, Zhou G, Kai G (2009) Molecular cloning, characterization and expression analysis of a new gene encoding 3-hydroxy-3-methylglutaryl coenzyme A reductase from Salvia miltiorrhiza. Acta Physiol Plant 31:565–572CrossRefGoogle Scholar
  31. Liao Y, Xu F, Huang X, Zhang W, Cheng H, Li L, Cheng S, Shen Y (2015) Promoter analysis and transcriptional profiling of Ginkgo biloba 3-hydroxy-3-methylglutaryl coenzyme A reductase (GbHMGR) gene in abiotic stress responses. Not Bot Horti Agrobo 43:25–34Google Scholar
  32. Liu W, Stewart CN Jr (2016) Plant synthetic promoters and transcription factors. Curr Opin Biotechnol 37:36–44CrossRefPubMedGoogle Scholar
  33. Liu Y, Xu QX, Wang XY, Liu CS, Chen HH (2012) Researches on the influence of 3-hydroxy-3-methylglutary-coenzyme A reductase gene polymorphism on catalytic efficiency of its encode enzyme in Glycyrrhiza uralensis. Zhongguo Zhong Yao Za Zhi 37:3784–3788PubMedGoogle Scholar
  34. Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) method. Methods 25:402–428CrossRefPubMedGoogle Scholar
  35. Lv DM, Zhang TT, Deng S, Zhang YH (2016) Functional analysis of the Malus domestica MdHMGR2 gene promoter in transgenic Arabidopsis thaliana. Biol Plant 60:667–676CrossRefGoogle Scholar
  36. Ma Y, Yuan L, Wu B, Li X, Chen S, Lu S (2012) Genome-wide identification and characterization of novel genes involved in terpenoid biosynthesis in Salvia miltiorrhiza. J Exp Bot 63:2809–2823CrossRefPubMedPubMedCentralGoogle Scholar
  37. Mao G, Seebeck T, Schrenker D, Yu O (2013) CYP709B3, a cytochrome P450 monooxygenase gene involved in salt tolerance in Arabidopsis thaliana. BMC Plant Biol 13:169CrossRefPubMedPubMedCentralGoogle Scholar
  38. Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15:473–497CrossRefGoogle Scholar
  39. Pedros R, Moya I, Goulas Y, Jacquemoud S (2008) Chlorophyll fluorescence spectrum inside a leaf. Photochem Photobiol Sci 7:498–502CrossRefPubMedGoogle Scholar
  40. Pfaffl MW, Tichopad A, Prgomet C, Neuvians TP (2004) Determination of stable housekeeping genes, differentially regulated target genes and sample integrity: bestKeeper–Excel-based tool using pair-wise correlations. Biotechnol Lett 26:509–515CrossRefPubMedGoogle Scholar
  41. Rajinikanth M, Harding SA, Tsai Ch-J (2007) The glycine decarboxylase complex multienzyme family in Populus. J Exp Bot 58:1761–1770CrossRefPubMedGoogle Scholar
  42. Saulierè J, Sureau A, Expert-Bezancon A, Marie J (2006) The polypyrimidine tract binding protein (PTB) represses splicing of exon 6B from the β-tropomyosin pre-mRNA by directly interfering with the binding of the U2AF65 subunit. Mol Cell Biol 26:8755–8769CrossRefPubMedPubMedCentralGoogle Scholar
  43. Schmittgen TD, Zakrajsek BA (2000) Effect of experimental treatment on housekeeping gene expression: validation by real-time, quantitative RT–PCR. J Biochem Biophys Methods 46:69–81CrossRefPubMedGoogle Scholar
  44. Shahmuradov IA, Gammerman AJ, Hancock JM, Bramley PM, Solovyev VV (2003) PlantProm: a database of plant promoter sequences. Nucleic Acids Res 31:114–117CrossRefPubMedPubMedCentralGoogle Scholar
  45. Shavrukov Y (2013) Salt stress or salt shock: which genes are we studying? J Exp Bot 64:119–127CrossRefPubMedGoogle Scholar
  46. Sunkar R, Zhou X, Zheng Y, Zhang W, Zhu J-K (2008) Identification of novel and candidate miRNAs in rice by high throughput sequencing. BMC Plant Biol 8:25CrossRefPubMedPubMedCentralGoogle Scholar
  47. Szymczyk P, Skała E, Grąbkowska R, Jeleń A, Żebrowska M, Balcerczak E (2016) Isolation and characterization of a copalyl diphosphate synthase gene promoter from Salvia miltiorrhiza. Acta Soc Bot Pol 85:3513CrossRefGoogle Scholar
  48. Tatematsu K, Ward S, Leyser O, Kamiya Y, Nambara E (2005) Identification of cis-elements that regulate gene expression during initiation of axillary bud outgrowth in Arabidopsis. Plant Physiol 138:757–766CrossRefPubMedPubMedCentralGoogle Scholar
  49. Teh KY, Abdullah JO (2016) An overview of 3-hydroxy-3-methylglutaryl CoA reductase (HMGR) in plants. Pertanika J Sch Res Rev 2:100–107Google Scholar
  50. Toufighi K, Brady SM, Austin R, Ly E, Provart NJ (2005) The botany array resource: e-Northerns, expression angling, and promoter analyses. Plant J 43:153–163CrossRefPubMedGoogle Scholar
  51. Usadel B, Obayashi T, Mutwil M, Giorgi FM, Bassel GW, Tanimoto N et al (2009) Co-expression tools for plant biology: opportunities for hypothesis generation and caveats. Plant Cell Environ 32:1633–1651CrossRefPubMedGoogle Scholar
  52. Vinces MD, Legendre M, Caldara M, Hagihara M, Verstrepen KJ (2009) Unstable tandem repeats in promoters confer transcriptional evolvability. Science 324:1213–1216CrossRefPubMedPubMedCentralGoogle Scholar
  53. Wang K (ed) (2006) Agrobacterium protocols. In: Methods in molecular biology, vol 343. Humana Press Inc, TotowaGoogle Scholar
  54. Wang H, Miyazaki S, Kawai K, Deyholos M, Galbraith DW, Bohnert HJ (2003) Temporal progression of gene expression responses to salt shock in maize roots. Plant Mol Biol 52:873–891CrossRefPubMedGoogle Scholar
  55. Weinhold A, Kallenbach M, Baldwin IT (2013) Progressive 35S promoter methylation increases rapidly during vegetative development in transgenic Nicotiana attenuata plants. BMC Plant Biol 13:99CrossRefPubMedPubMedCentralGoogle Scholar
  56. Wu SJ, Shi M, Wu JY (2009a) Cloning and characterization of the 1-deoxy-D-xylulose 5-phosphate reductoisomerase gene for diterpenoid tanshinone biosynthesis in Salvia miltiorrhiza (Chinese sage) hairy roots. Biotechnol Appl Biochem 52:89–95CrossRefPubMedGoogle Scholar
  57. Wu L, Zhang Q, Zhou H, Ni F, Wu X, Qia Z (2009b) Rice MicroRNA effector complexes and targets. Plant Cell 21:3421–3435CrossRefPubMedPubMedCentralGoogle Scholar
  58. Xu X, Jiang Q, Ma X, Ying Q, Shen B, Qian Y, Song H, Wang H (2014) Deep sequencing identifies tissue-specific microRNAs and their target genes involving in the biosynthesis of Tanshinones in Salvia miltiorrhiza. PLoS ONE 9:e111679CrossRefPubMedPubMedCentralGoogle Scholar
  59. Yan Y, Wang Z (2007) Genetic transformation of the medicinal plant Salvia miltiorrhiza by Agrobacterium tumefaciens-mediated method. Plant Cell Organ Tiss Cult 88:175–184CrossRefGoogle Scholar
  60. Yang Y, Hou S, Cui G, Chen S, Wei J, Huang L (2010) Characterization of reference genes for quantitative real time PCR analysis in various tissues of Salvia miltiorrhiza. Mol Biol Rep 37:507–513CrossRefPubMedGoogle Scholar
  61. Zhang Y, Zhu Y, Peng Y, Yan D, Li Q, Wang J, Wang L, He Z (2008) Gibberellin homeostasis and plant height control by EUI and a role for gibberellin in root gravity responses in rice. Cell Res 18:412–421CrossRefPubMedGoogle Scholar
  62. Zhang X, Guo J, Shen Y, Huang L (2012) Cloning and expression analysis of a new 3-hydroxy-3-methylglutaryl coenzyme A reductase gene from Salvia miltiorrhiza (SmHMGR3). Zhongguo Zhong Yao Za Zhi 37:2378–2382PubMedGoogle Scholar
  63. Zhou L, Zuo Z, Chow MS (2005) Danshen: an overview of its chemistry, pharmacology, pharmacokinetics, and clinical use. J Clin Pharmacol 45:1345–1359CrossRefPubMedGoogle Scholar

Copyright information

© Society for Plant Biochemistry and Biotechnology 2017

Authors and Affiliations

  • Piotr Szymczyk
    • 1
  • Renata Grąbkowska
    • 2
  • Ewa Skała
    • 2
  • Marta Żebrowska
    • 3
  • Ewa Balcerczak
    • 3
  • Agnieszka Jeleń
    • 3
  1. 1.Department of Pharmaceutical BiotechnologyMedical University of ŁódźLodzPoland
  2. 2.Department of Biology and Pharmaceutical BotanyMedical University of ŁódźLodzPoland
  3. 3.Department of Pharmaceutical Biochemistry and Molecular DiagnosticsMedical University of ŁódźLodzPoland

Personalised recommendations