The dirigent multigene family in Isatis indigotica: gene discovery and differential transcript abundance

BMC Genomics

Wansheng Chen³  Lei Zhang¹  Peng Di³  Ying Xiao²  Junfeng Chen³  Qing Li³ 

[1] Department of Pharmaceutical Botany, School of Pharmacy, Second Military Medical University, No. 325 Guohe Road, Yangpu District, Shanghai 200433, China;The MOE Key Laboratory of Standardization of Chinese Medicines, Institute of Chinese Materia Medica,Shanghai University of Traditional Chinese Medicine, Shanghai 201210, China;Department of Pharmacy, Shanghai Changzheng Hospital, Second Military Medical University, No. 415 Fengyang Road, Huangpu District, Shanghai 200003, China

关键词: Transcript abundance;    Phylogenetic analysis;    Tertiary structures;    Secondary structures;    Bioinformatics;    Isatis indigotica;    Dirigent and dirigent-like proteins;   

Others  :  1217210
DOI  :  10.1186/1471-2164-15-388

 received in 2013-12-09, accepted in 2014-05-14,  发布年份 2014

【 摘 要 】

Background

Isatis indigotica Fort. is one of the most commonly used traditional Chinese medicines. Its antiviral compound is a kind of lignan, which is formed with the action of dirigent proteins (DIR). DIR proteins are members of a large family of proteins which impart stereoselectivity on the phenoxy radical-coupling reaction, yielding optically active lignans from two molecules of E-coniferyl alcohol. They exist in almost every vascular plant. However, the DIR and DIR-like protein gene family in I. indigotica has not been analyzed in detail yet. This study focuses on discovery and analysis of this protein gene family in I. indigotica for the first time.

Results

Analysis of transcription profiling database from I. indigotica revealed a family of 19 full-length unique DIR and DIR-like proteins. Sequence analysis found that I. indigotica DIR and DIR-like proteins (IiDIR) were all-beta strand proteins, with a signal peptide at the N-terminus. Phylogenetic analysis of the 19 proteins indicated that the IiDIR genes cluster into three distinct subfamilies, DIR-a, DIR-b/d, and DIR-e, of a larger plant DIR and DIR-like gene family. Gene-specific primers were designed for 19 unique IiDIRs and were used to evaluate patterns of constitutive expression in different organs. It showed that most IiDIR genes were expressed comparatively higher in roots and flowers than stems and leaves.

Conclusions

New DIR and DIR-like proteins were discovered from the transcription profiling database of I. indigotica through bioinformatics methods for the first time. Sequence characteristics and transcript abundance of these new genes were analyzed. This study will provide basic data necessary for further studies.

【 授权许可】

   
2014 Li et al.; licensee BioMed Central Ltd.

【 图 表 】

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【 参考文献 】

• [1]Committee NP: Chinese Pharmacopoeia. Beijing: China Medical Science Press; 2010:191.
• [2]Tietao D: Discussion on treatment of SARS by TCM. Tianjin J Traditional Chin Med 2003, 3:001.
• [3]Li B: Active Ingredients and Quality Evaluation of Isatis Indigotica. Shanghai: Second Military Medical University; 2003.
• [4]Saleem M, Kim HJ, Ali MS, Lee YS: An update on bioactive plant lignans. Nat Prod Rep 2005, 22(6):696-716.
• [5]Adlercreutz H, Mazur W: Phyto-oestrogens and Western diseases. Ann Med 1997, 29(2):95-120.
• [6]Arts IC, van de Putte B, Hollman PC: Catechin contents of foods commonly consumed in The Netherlands. 1. Fruits, vegetables, staple foods, and processed foods. J Agric Food Chem 2000, 48(5):1746-1751.
• [7]Adlercreutz H, Mousavi Y, Clark J, Höckerstedt K, Hämäläinen E, Wähälä K, Mäkelä T, Hase T: Dietary phytoestrogens and cancer: in vitro and in vivo studies. J Steroid Biochem Mol Biol 1992, 41(3):331-337.
• [8]Raffaelli B, Hoikkala A, Leppälä E, Wähälä K: Enterolignans. J Chromatogr B 2002, 777(1):29-43.
• [9]Arts IC, Hollman PC: Polyphenols and disease risk in epidemiologic studies. Am J Clin Nutr 2005, 81(1):317S-325S.
• [10]Burlat V, Kwon M, Davin LB, Lewis NG: Dirigent proteins and dirigent sites in lignifying tissues. Phytochemistry 2001, 57(6):883-897.
• [11]Pickel B, Pfannstiel J, Steudle A, Lehmann A, Gerken U, Pleiss J, Schaller A: A model of dirigent proteins derived from structural and functional similarities with allene oxide cyclase and lipocalins. FEBS J 2012, 279(11):1980-1993.
• [12]Davin LB, Wang H-B, Crowell AL, Bedgar DL, Martin DM, Sarkanen S, Lewis NG: Stereoselective bimolecular phenoxy radical coupling by an auxiliary (dirigent) protein without an active center. Science 1997, 275(5298):362-367.
• [13]Halls SC, Lewis NG: Secondary and quaternary structures of the (+)-pinoresinol-forming dirigent protein. Biochemistry 2002, 41(30):9455-9461.
• [14]Halls SC, Davin LB, Kramer DM, Lewis NG: Kinetic study of coniferyl alcohol radical binding to the (+)-pinoresinol forming dirigent protein. Biochemistry 2004, 43(9):2587-2595.
• [15]Davin LB, Lewis NG: Dirigent proteins and dirigent sites explain the mystery of specificity of radical precursor coupling in lignan and lignin biosynthesis. Plant Physiol 2000, 123(2):453-462.
• [16]Ralph S, Park J-Y, Bohlmann J, Mansfield SD: Dirigent proteins in conifer defense: gene discovery, phylogeny, and differential wound-and insect-induced expression of a family of DIR and DIR-like genes in spruce (Picea spp.). Plant Mol Biol 2006, 60(1):21-40.
• [17]Ralph SG, Jancsik S, Bohlmann J: Dirigent proteins in conifer defense II: extended gene discovery, phylogeny, and constitutive and stress-induced gene expression in spruce (Picea spp.). Phytochemistry 2007, 68(14):1975-1991.
• [18]Chen J, Dong X, Li Q, Zhou X, Gao S, Chen R, Sun L, Zhang L, Chen W: Biosynthesis of the active compounds of Isatis indigotica based on transcriptome sequencing and metabolites profiling. BMC Genomics 2013, 14(1):857. BioMed Central Full Text
• [19]Thamil Arasan SK, Park J-I, Ahmed NU, Jung H-J, Hur Y, Kang K-K, Lim Y-P, Nou I-S: Characterization and expression analysis of dirigent family genes related to stresses in Brassica. Plant Physiol Biochem 2013, 67:144-153.
• [20]Letunic I, Doerks T, Bork P: SMART 7: recent updates to the protein domain annotation resource. Nucleic Acids Res 2012, 40(D1):D302-D305.
• [21]Emanuelsson O, Nielsen H, Brunak S, Von Heijne G: Predicting subcellular localization of proteins based on their N-terminal amino acid sequence. J Mol Biol 2000, 300(4):1005-1016.
• [22]Horton P, Park K-J, Obayashi T, Fujita N, Harada H, Adams-Collier C, Nakai K: WoLF PSORT: protein localization predictor. Nucleic Acids Res 2007, 35(suppl 2):W585-W587.
• [23]Gupta R, Jung E, Brunak S: Prediction of N-glycosylation sites in human proteins. preparation 2004. 2004
• [24]Gasteiger E, Hoogland C, Gattiker A, Duvaud S, Wilkins MR, Appel RD, Bairoch A: Protein identification and analysis tools on the ExPASy server. In The proteomics protocols handbook. Edited by Walker JM. Totowa, N.J: Humana Press; 2005:571-607.
• [25]Petersen B, Petersen TN, Andersen P, Nielsen M, Lundegaard C: A generic method for assignment of reliability scores applied to solvent accessibility predictions. BMC Struct Biol 2009, 9(1):51. BioMed Central Full Text
• [26]Kelley LA, Sternberg MJ: Protein structure prediction on the Web: a case study using the Phyre server. Nat Protoc 2009, 4(3):363-371.
• [27]Pickel B, Constantin MA, Pfannstiel J, Conrad J, Beifuss U, Schaller A: An enantiocomplementary dirigent protein for the enantioselective laccase‒catalyzed oxidative coupling of phenols. Angew Chem Int Ed 2010, 49(1):202-204.
• [28]Davin LB, Lewis NG: Dirigent phenoxy radical coupling: advances and challenges. Curr Opin Biotechnol 2005, 16(4):398-406.
• [29]Kim MK, Jeon J-H, Fujita M, Davin LB, Lewis NG: The western red cedar (Thuja plicata) 8-8’ DIRIGENT family displays diverse expression patterns and conserved monolignol coupling specificity. Plant Mol Biol 2002, 49(2):199-214.
• [30]Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG: The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 1997, 25(24):4876-4882.
• [31]Kim MK, Jeon J-H, Davin LB, Lewis NG: Monolignol radical–radical coupling networks in western red cedar and Arabidopsis and their evolutionary implications. Phytochemistry 2002, 61(3):311-322.
• [32]Gang DR, Costa MA, Fujita M, Dinkova-Kostova AT, Wang H-B, Burlat V, Martin W, Sarkanen S, Davin LB, Lewis NG: Regiochemical control of monolignol radical coupling: a new paradigm for lignin and lignan biosynthesis. Chem Biol 1999, 6(3):143-151.
• [33]Letunic I, Bork P: Interactive Tree Of Life (iTOL): an online tool for phylogenetic tree display and annotation. Bioinformatics 2007, 23(1):127-128.
• [34]Saeed A, Sharov V, White J, Li J, Liang W, Bhagabati N, Braisted J, Klapa M, Currier T, Thiagarajan M, Sturn A, Snuffin M, Rezantsev A, Popov D, Ryltsov A, Kostukovich E, Borisovsky I, Liu Z, Vinsavich A, Trush V, Quackenbush J: TM4: a free, open-source system for microarray data management and analysis. Biotechniques 2003, 34(2):374-378.