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Identification, validation, and expression of ABC transporters in Podophyllum hexandrum and their role in podophyllotoxin biosynthesis

  • Original papers
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Biologia Plantarum

Abstract

Podophyllum hexandrum Royle is an important medicinal herb of North-Western Himalayas, and podophyllotoxin, being its major metabolite, has been used extensively in the preparation of several anticancer drugs. Podophyllotoxin accumulates in rhizomes; however, no information exists on the role of ATP-binding cassette (ABC) transporters vis-à-vis podophyllotoxin content. The present study reports identification, validation, and expression analysis of ABC transporter genes from P. hexandrum. Total 252 ABC transporter genes were identified as unigenes out of which 22 were further validated using real time qPCR in different tissues of varying podophyllotoxin content. Differential expression analysis and Pearson’s correlation coefficient revealed two candidate genes PhABC6 and PhABCIII having a positive correlation with the podophyllotoxin content. PhABCIV showed the highest expression in rhizomes (20.53-folds compared to shoots) suggesting its possible role in transport and accumulation of podophyllotoxin.

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Abbreviations

ABC:

ATP-binding cassette

miRNA:

microRNA

FPKM:

fragments per kilobase of transcript per million mapped reads

NGS:

next-generation sequencing

PERL:

Practical Extraction and Report Language

Pfam:

protein family database

qPCR:

quantitative polymerase chain reaction

RSEM:

RNA sequences by expectation maximization

TFs:

transcription factors

References

  • Alejandro, S., Lee, Y., Tohge, T., Sudre, D., Osorio, S., Park, J., Bovet, L., Lee, Y., Geldner, N., Fernie, A.R., Martinoia, E.: AtABCG29 is a monolignol transporter involved in lignin biosynthesis. — Curr. Biol. 22: 1207–1212, 2012.

    Article  CAS  PubMed  Google Scholar 

  • Andolfo, G., Ruocco, M., Donato, A.D., Frusciante, L., Lorito, M., Scala, F., Ercolano M.R.: Genetic variability and evolutionary diversification of membrane ABC transporters in plants. — BMC Plant Biol. 15: 5, 2015.

    Article  Google Scholar 

  • Armisen, D., Lecharny, A., Aubourg, S.: Unique genes in plants: specificities and conserved features throughout evolution. — BMC Evol. Biol. 8: 280, 2008.

    Article  PubMed  PubMed Central  Google Scholar 

  • Badri, D.V., Chaparro, J.M., Manter, D.K., Martinoia, E., Vivanco, J.M.: Influence of ATP-binding cassette transporters in root exudation of phytoalexins, signals, and in disease resistance. — Front. Plant Sci. 3: 149, 2012.

    Article  PubMed  PubMed Central  Google Scholar 

  • Badri, D.V., Quintana, N, El Kassis, E.G., Kim, H.K., Choi, Y.H., Sugiyama, A., Verpoorte, R., Martinoia, E., Manter, D.K., Vivanco, J.M.: An ABC transporter mutation alters root exudation of phytochemicals that provoke an overhaul of natural soil microbiota. — Plant Physiol. 151: 2006–2017, 2009.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bhattacharyya, D., Sinha, R., Datta, R., Hazra, S., Chattopadhyay, S.: De novo transcriptome analysis using 454 pyrosequencing of the Himalayan may apple, Podophyllum hexandrum. — BMC Genomics 14: 748, 2013.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Canel, C., Dayan, F.E., Ganzera, M., Khan, I.A., Rimando, A., Burandt, C.L., Moraes, R.M.: High yield of podophyllotoxin from leaves of Podophyllum peltatum by in situ conversion of podophyllotoxin 4-O-β-D glucopyranoside. — Planta med. 67: 97–99, 2001.

    Article  CAS  PubMed  Google Scholar 

  • Canel, C., Moreas, R.M., Dyan, F.E., Ferreira, D.: Molecules of interest: podophyllotoxin. — Phytochemistry 54: 115–120, 2000.

    Article  CAS  PubMed  Google Scholar 

  • Francisco, R.M., Regalado, A., Ageorges, A., Burla, B.J., Bassin, B., Eisenach, C., Zarrouk, O.V.S., Marlin, T., Chaves, M.M., Martinoia, E., Nagy, R.: ABCC1, an ATP binding cassette protein from grape berry, transports anthocyanidin 3-O-glucosides. — Plant Cell Online 25: 1840–1854, 2013.

    Article  CAS  Google Scholar 

  • Grec, S., Vanham, D., Ribaucourt, J.C., Purnelle, B., Boutry, M.: Identification of regulatory sequence elements within the transcription promoter region of PhABCV, a gene encoding a plant ABC transporter induced by diterpenes. — Plant J. 35: 237–250, 2003.

    Article  CAS  PubMed  Google Scholar 

  • Holland, I.B., Cole, S.P.C., Kuchler, K., Higgins C.F.: ABC Proteins: From Bacteria to Man. — Academic Press, San Diego 2003.

    Google Scholar 

  • Huber, S.C.: Biochemical mechanism for regulation of sucrose accumulation in leaves during photosynthesis. — Plant Physiol. 91: 656–662, 1989.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Jeong, C.B., Kim, B.M., Lee, J.S.: Genome-wide identification of whole ATP-binding cassette (ABC) transporters in the intertidal copepod Tigriopus japonicus. — BMC Genomics 15: 651, 2014.

    Article  PubMed  PubMed Central  Google Scholar 

  • Kang, J., Park, J., Choi, H., Burla, B., Kretzschmar, T., Lee, Y., Martinoia, E.: Plant ABC Transporters. — The Arabidopsis Book/American Society of Plant Biologists. 9: e0153, 2011.

    Google Scholar 

  • Kuhlmann, S., Kranz, K., Lücking, B., Alfermann, A.W., Petersen, M.: Aspects of cytotoxic lignan biosynthesis in suspension cultures of Linum nodiflorum. — Phytochem. Rev. 1: 37–43, 2002.

    Article  CAS  Google Scholar 

  • Kumar, P., Pal, T., Sharma, N., Kumar, V., Sood, H., Chauhan, R.S.: Expression analysis of biosynthetic pathway genes vis-à-vis podophyllotoxin content in Podophyllum hexandrum Royle. — Protoplasma 252: 1252–1262, 2015.

    Google Scholar 

  • Kunze, R., Frommer, W.B., Flügge, U.I.: Metabolic engineering of plants: the role of membrane transport. — Metabolic Engn. 4: 57–66, 2002.

    Article  CAS  Google Scholar 

  • Marques, J.V., Kim, K.W., Lee, C., Costa, M.A., May, G.D., Crow, J.A., Davin, L.B., Lewis N.G.: Next generation sequencing in predicting gene function in podophyllotoxin biosynthesis. — J. biol. Chem. 288: 466–479, 2013.

    Article  CAS  PubMed  Google Scholar 

  • Martinoia, E., Grill, E., Tommasini, R., Kreuz, K., Amrhein, N.: ATP-dependent glutathione S-conjugate 'export' pump in the vacuolar membrane of plants. — Nature 364: 247–249, 1993.

    Article  CAS  Google Scholar 

  • Martinoia, E., Klein, M., Geisler, M., Bovet, L., Forestier, C., Kolukisaoglu, U., Muller-Rober, B., Schulz, B.: Multifunctionality of plant ABC transporters — more than just detoxifiers. — Planta 214: 345–355, 2002.

    Article  CAS  PubMed  Google Scholar 

  • Moraes, R.M.: The lignans of Podophyllum. — Stud. natur. Products Chem. 2: 149–182, 2002.

    Article  Google Scholar 

  • Pal, T., Malhotra, N., Chanumolu, S.K., Chauhan, R.S.: Next-generation sequencing (NGS) transcriptomes reveal association of multiple genes and pathways contributing to secondary metabolites accumulation in tuberous roots of Aconitum heterophyllum Wall. — Planta 242: 239–258, 2015.

    Article  CAS  PubMed  Google Scholar 

  • Pandit, S., Shitiz, K., Sood, H., Naik, P.K., Chauhan, R.S.: Expression pattern of fifteen genes of non-mevalonate (MEP) and mevalonate (MVA) pathways in different tissues of endangered medicinal herb Picrorhiza kurroa with respect to picrosides content. — Mol. Biol. Rep. 40: 1053–1063, 2013.

    Article  CAS  PubMed  Google Scholar 

  • Raichaudhuri, A., Peng, M., Naponelli, V., Chen, S., Sánchez-Fernández, R., Gu, H., Gregory, J.F., Hanson, A.D., Rea, P.A.: Plant vacuolar ATP-binding cassette transporters that translocate folates and anti-folates in vitro and contribute to anti-folate tolerance in vivo. — J. biol. Chem. 284: 8449–8460, 2009.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sánchez-Fernández, R., Davies, T.G., Coleman, J.O., Rea, P.A.: The Arabidopsis thaliana ABC protein superfamily, a complete inventory. — J. biol. Chem. 276: 30231–30244, 2001.

    Article  PubMed  Google Scholar 

  • Sibout, R., Höfte, H.: Plant cell biology: the ABC of monolignol transport. — Curr. Biol. 22: 533–535, 2012.

    Article  Google Scholar 

  • Song, W.Y., Park, J., Mendoza-Cózatl, D.G., Suter-Grotemeyer, M., Shim, D., Hörtensteiner, S., Geisler, M., Weder, B., Rea, P.A., Rentsch, D., Schroeder, J.I., Lee, Y., Martinoia, E.: Arsenic tolerance in Arabidopsis is mediated by two ABCC-type phytochelatin transporters. — Proc. nat. Acad. Sci. USA 107: 21187–21192, 2010.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Tabata, M.: The mechanism of shikonin biosynthesis in Lithospermum cell cultures. — Plant Tissue Cult. Lett. 13: 117–125, 1996.

    Article  CAS  Google Scholar 

  • Theodolou, F.L.: Plant ABC transporters. — Biochim. biophys. Acta 1465: 79–103, 2000.

    Article  Google Scholar 

  • Tamura, K., Stecher, G., Peterson, D., Filipski, A., Kumar, S.: MEGA6: molecular evolutionary genetics analysis version 6.0. — Mol. Biol. Evolut. 30: 2725–2729, 2013.

    Article  CAS  Google Scholar 

  • Van Uden, W., Pras, N., Visser, J.F., Malingre, T.M.: Detection and identification of podophyllotoxin produced by cell cultures derived from Podophyllum hexandrum Royle. — Plant Cell Rep. 8: 165–168, 1989.

    Article  PubMed  Google Scholar 

  • Yazaki, K.: Transporters of secondary metabolites. — Curr. Opin. Plant Biol. 8: 301–307, 2005.

    Article  CAS  PubMed  Google Scholar 

  • Yazaki, K., Sugiyama, A., Morita, M., Shitan, N.: Secondary transport as an efficient membrane transport mechanism for plant secondary metabolites. — Phytochem. Rev. 7: 513–524, 2008.

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Biotechnology and Bioinformatics, Jaypee University of Information Technology, Waknaghat, Solan, 173234, Himachal Pradesh, India

    P. Kumar, R. Sharma, V. Jaiswal & R. S. Chauhan

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  1. P. Kumar
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  2. R. Sharma
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  3. V. Jaiswal
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  4. R. S. Chauhan
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Corresponding author

Correspondence to R. S. Chauhan.

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Acknowledgement

The authors are thankful to the Jaypee University of Information Technology for providing the necessary research facilities. The authors are also thankful to the Department of Biotechnology, the Ministry of Science & Technology, Govt. of India for a grant to RSC, and to the Himalayan Forest Research Institute, Panthaghati, Shimla, India for providing the plant material of P. hexandrum. The work presented here was carried out in collaboration of all authors.

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Kumar, P., Sharma, R., Jaiswal, V. et al. Identification, validation, and expression of ABC transporters in Podophyllum hexandrum and their role in podophyllotoxin biosynthesis. Biol Plant 60, 452–458 (2016). https://doi.org/10.1007/s10535-016-0611-9

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  • DOI: https://doi.org/10.1007/s10535-016-0611-9

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