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Cloning and functional characterization of PjPORB, a member of the POR gene family in Pseudosasa japonica cv. Akebonosuji

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Abstract

In angiosperms, NADPH: protochlorophyllide oxidoreductase (POR) catalyzes the photoreduction of protochlorophyllide (Pchlide) to chlorophyllide (Chlide), the only light-dependent step in chlorophyll biosynthesis. There is a wide variety of gene organization and light- and development-dependent regulatory mechanisms for the POR genes. In this study, a POR homologue, designated PjPORB, was isolated from Pseudosasa japonica cv. Akebonosuji, which is a cultivar of high ornamental value in landscape due to its unique green-white striped leaf phenotype. The full-length PjPORB cDNA was 1567 bp long with a 1185 bp ORF that encoded 394 amino acids. Multiple amino acid sequence alignment showed that the putative PjPORB shared a high similarity to POR homologues from other plant species, and that the cofactor (NADPH)-binding motif and active site motif in particular were highly conserved among all the PORs. Further, PjPORB was overexpressed in Arabidopsis thaliana; and the Pchlide contents of the genetically modified plants were reduced to a larger extent than that of wild type plants, thereby indicating its important role in Pchlide photoreduction. In real-time quantitative RT-PCR analysis, PjPORB was expressed at higher levels in leaf samples than in culms and roots, and showed a first increasing-then decreasing expression pattern during the development of white, stripe, and green leaf samples, thus indicating its essential role in leaf development. Although the mRNA levels for PjPORB did not differ considerably among white, stripe, and green leaf samples within the two almost same earlier developmental stages, its expression levels in the late leaf developmental stage fell into two distinct classes: high expression levels in strip and green leaf samples, and low expression levels in white leaf. Collectively, these expression data suggested that PjPORB may be involved in the leaf color variation for P. japonica cv. Akebonosuji.

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References

  • Alawady AE, Grimm B (2005) Tobacco Mg protoporphyrin IX methyltransferase is involved in inverse activation of Mg porphyrin and protoheme synthesis. Plant J 41:282–290

    Article  CAS  PubMed  Google Scholar 

  • Armstrong GA, Runge S, Frick G, Sperling U, Apel K (1995) Identification of NADPH: protochlorophyllide oxidoreductases A and B: a branched pathway for light-dependent chlorophyll biosynthesis in Arabidopsis thaliana. Plant Physiol 108:1505–1517

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Benli M, Schulz R, Apel K (1991) Effect of light on the NADPH-protochlorophyllide oxidoreductase of Arabidopsis thaliana. Plant Mol Biol 16:615–625

    Article  CAS  PubMed  Google Scholar 

  • Clough SJ, Bent AF (1998) Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J 16:735–743

    Article  CAS  PubMed  Google Scholar 

  • Darrah PM, Kay S, Teakle GR, Griffiths WT (1990) Cloning and sequencing of protochlorophyllide reductase. Biochem J 265:789–798

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fan CJ, Ma JM, Guo QR, Li XT, Wang H, Lu MZ (2013) Selection of reference genes for quantitative real-time PCR in bamboo (Phyllostachys edulis). PLoS ONE 8:e56573

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Forreiter C, van Cleve B, Schmidt A, Apel K (1991) Evidence for a general light-dependent negative control of NADPH-protochlorophyllide oxidoreductase in angiosperms. Planta 183:126–132

    Article  CAS  PubMed  Google Scholar 

  • Frick G, Su Q, Apel K, Armstrong GA (2003) An Arabidopsis porB porC double mutant lacking light-dependent NADPH: protochlorophyllide oxidoreductases B and C is highly chlorophyll-deficient and developmentally arrested. Plant J 35:141–153

    Article  CAS  PubMed  Google Scholar 

  • Griffiths WT (1978) Reconstitution of chlorophyllide formation by isolated etioplast membranes. Biochem J 174:681–692

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hodgins R, Van Huystee R (1986) Rapid simultaneous estimation of protoporphyrin and Mg-porphyrins in higher plants. J Plant Physiol 125:311–323

    Article  CAS  Google Scholar 

  • Holtorf H, Reinbothe S, Reinbothe C, Bereza B, Apel K (1995) Two routes of chlorophyllide synthesis that are differentially regulated by light in barley (Hordeum vulgare L.). P Natl Acad Sci USA 92:3254–3258

    Article  CAS  Google Scholar 

  • Hopkins W (1982) Formation of chloroplast pigments in a temperature-sensitive, virescent mutant of maize. Can J Bot 60:737–740

    Article  CAS  Google Scholar 

  • Hopkins W, Elfman B (1984) Temperature-induced chloroplast ribosome deficiency in virescent maize. J Hered 75:207–211

    CAS  Google Scholar 

  • Iwamoto K, Fukuda H, Sugiyama M (2001) Elimination of POR expression correlates with red leaf formation in Amaranthus tricolor. Plant J 27:275–284

    Article  CAS  PubMed  Google Scholar 

  • Kusumi K, Chono Y, Shimada H, Gotoh E, Tsuyama M, Iba K (2010) Chloroplast biogenesis during the early stage of leaf development in rice. Plant Biotechnol 27:85–90

    Article  CAS  Google Scholar 

  • Lang A (1952) Physiology of flowering. Annu Rev Plant Physiol 3:265–306

    Article  Google Scholar 

  • Lebedev N, Timko MP (1998) Protochlorophyllide photoreduction. Photosynth Res 58:5–23

    Article  CAS  Google Scholar 

  • Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25:402–408

    Article  CAS  PubMed  Google Scholar 

  • Masuda T, Takamiya K (2004) Novel Insights into the enzymology, regulation and physiological functions of light-dependent protochlorophyllide oxidoreductase in angiosperms. Photosynth Res 81:1–29

    Article  CAS  PubMed  Google Scholar 

  • Masuda T, Fusada N, Shiraishi T, Kuroda H, Awai K, Shimada H, Ohta H, Takamiya K (2002) Identification of two differentially regulated isoforms of protochlorophyllide oxidoreductase (POR) from tobacco revealed a wide variety of light- and development-dependent regulations of POR gene expression among angiosperms. Photosynth Res 74:165–172

    Article  CAS  PubMed  Google Scholar 

  • Millerd A, McWilliam J (1968) Studies on a maize mutant sensitive to low temperature I. Influence of temperature and light on the production of chloroplast pigments. Plant Physiol 43:1967–1972

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  • Oosawa N, Masuda T, Awai K, Fusada N, Shimada H, Ohta H, Takamiya K (2000) Identification and light-induced expression of a novel gene of NADPH-protochlorophyllide oxidoreductase isoform in Arabidopsis thaliana. FEBS Lett 474:133–136

    Article  CAS  PubMed  Google Scholar 

  • Paddock T, Lima D, Mason ME, Apel K, Armstrong GA (2012) Arabidopsis light-dependent protochlorophyllide oxidoreductase A (PORA) is essential for normal plant growth and development. Plant Mol Biol 78:447–460

    Article  CAS  PubMed  Google Scholar 

  • Pattanayak GK, Tripathy BC (2011) Overexpression of protochlorophyllide oxidoreductase C regulates oxidative stress in Arabidopsis. PLoS ONE 6:e26532

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Peng ZH, Lu Y, Li LB, Zhao Q, Feng Q, Gao ZM, Lu HY, Hu T, Yao N, Liu KY (2013) The draft genome of the fast-growing non-timber forest species moso bamboo (Phyllostachys heterocycla). Nat Genet 45:456–461

    Article  CAS  PubMed  Google Scholar 

  • Pogson BJ, Albrecht V (2011) Genetic dissection of chloroplast biogenesis and development: an overview. Plant Physiol 155:1545–1551

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rebeiz CA, Montazer-Zouhoor A, Mayasich JM, Tripathy BC, Wu SM, Rebeiz CC, Friedmann HC (1988) Photodynamic herbicides: recent developments and molecular basis of selectivity. Crit Rev Plant Sci 6:385–436

    Article  CAS  Google Scholar 

  • Richter A, Peter E, Pörs Y, Lorenzen S, Grimm B, Czarnecki O (2010) Rapid dark repression of 5-aminolevulinic acid synthesis in green barley leaves. Plant Cell Physiol 51:670–681

    Article  CAS  PubMed  Google Scholar 

  • Sakuraba Y, Rahman ML, Cho SH, Kim YS, Koh HJ, Yoo SC, Paek NC (2013) The rice faded green leaf locus encodes protochlorophyllide oxidoreductase B and is essential for chlorophyll synthesis under high light conditions. Plant J 74:122–133

    Article  CAS  PubMed  Google Scholar 

  • Schulz R, Steinmüller K, Klaas M, Forreiter C, Rasmussen S, Hiller C, Apel K (1989) Nucleotide sequence of a cDNA coding for the NADPH-protochlorophyllide oxidoreductase (PCR) of barley (Hordeum vulgare L.) and its expression in Escherichia coli. Mol Gen Genet 217:355–361

    Article  CAS  PubMed  Google Scholar 

  • Shalygo N, Czarnecki O, Peter E, Grimm B (2009) Expression of chlorophyll synthase is also involved in feedback-control of chlorophyll biosynthesis. Plant Mol Biol 71:425–436

    Article  CAS  PubMed  Google Scholar 

  • Spano AJ, He Z, Michel H, Hunt DF, Timko MP (1992) Molecular cloning, nuclear gene structure, and developmental expression of NADPH: protochlorophyllide oxidoreductase in pea (Pisum sativum L.). Plant Mol Biol 18:967–972

    Article  CAS  PubMed  Google Scholar 

  • Stobart A, Ameen-Bukhari I (1984) Regulation of delta-aminolaevulinic acid synthesis and protochlorophyllide regeneration in the leaves of dark-grown barley (Hordeum vulgare) seedlings. Biochem J 222:419–426

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Su Q, Frick G, Armstrong G, Apel K (2001) POR C of Arabidopsis thaliana: a third light- and NADPH-dependent protochlorophyllide oxidoreductase that is differentially regulated by light. Plant Mol Biol 47:805–813

    Article  CAS  PubMed  Google Scholar 

  • Talaat NB (2013) RNAi based simultaneous silencing of all forms of light-dependent NADPH: protochlorophyllide oxidoreductase genes result in the accumulation of protochlorophyllide in tobacco (Nicotiana tabacum). Plant Physiol Biochem 71:31–36

    Article  CAS  PubMed  Google Scholar 

  • Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Teakle GR, Griffiths WT (1993) Cloning, characterization and import studies on protochlorophyllide reductase from wheat (Triticum aestivum). Biochem J 296:225–230

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tripathy BC, Chakraborty N (1991) 5-Aminolevulinic acid induced photodynamic damage of the photosynthetic electron transport chain of cucumber (Cucumis sativus L.) cotyledons. Plant Physiol 96:761–767

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Waters MT, Langdale JA (2009) The making of a chloroplast. EMBO J 28:2861–2873

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wu W, Elsheery N, Wei Q, Zhang L, Huang J (2011) Defective etioplasts observed in variegation mutants may reveal the light-independent regulation of white/yellow sectors of Arabidopsis leaves. J Integr Plant Biol 53:846–857

    Article  CAS  PubMed  Google Scholar 

  • Yoshida K, Chen R-M, Tanaka A, Teramoto H, Tanaka R, Timko MP, Tsuji H (1995) Correlated changes in the activity, amount of protein, and abundance of transcript of NADPH: protochlorophyllide oxidoreductase and chlorophyll accumulation during greening of cucumber cotyledons. Plant Physiol 109:231–238

    CAS  PubMed  PubMed Central  Google Scholar 

  • Zavaleta-Mancera H, Franklin K, Ougham H, Thomas H, Scott I (1999) Regreening of senescent Nicotiana leaves I. Reappearance of NADPH-protochlorophyllide oxidoreductase and light-harvesting chlorophyll a/b-binding protein. J Exp Bot 50:1677–1682

    CAS  Google Scholar 

  • Zhao HS, Peng ZH, Fei BH, Li LB, Hu T, Gao ZM, Jiang ZH (2014) BambooGDB: a bamboo genome database with functional annotation and an analysis platform. Database (Oxford) 2014:bau006

    Article  Google Scholar 

  • Zheng BS (2006) Modern plant physiological and biochemical research technology. China Meteorological Press, Beijing

    Google Scholar 

Download references

Acknowledgments

This work was supported by the grant from the National Natural Science Foundation of China (31170565, 31270645 and 31470615), the Program of Natural Science Foundation of Zhejiang Province (LR12C16001 and LY12C16002).

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  1. The Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Linan, Zhejiang Province, People’s Republic of China

    Keyi Jiang & Mingbing Zhou

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  1. Keyi Jiang
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Correspondence to Mingbing Zhou.

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Jiang, K., Zhou, M. Cloning and functional characterization of PjPORB, a member of the POR gene family in Pseudosasa japonica cv. Akebonosuji. Plant Growth Regul 79, 95–106 (2016). https://doi.org/10.1007/s10725-015-0115-1

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