Journal of Plant Research

, Volume 129, Issue 4, pp 737–748 | Cite as

Gene characterization and transcription analysis of two new ammonium transporters in pear rootstock (Pyrus betulaefolia)

  • Hui Li
  • Jin-Long Han
  • You-Hong Chang
  • Jing Lin
  • Qing-Song Yang
Regular Paper

Abstract

Ammonium is the primarily nitrogen source for plant growth, but the molecular basis of ammonium acquisition in fruit species remains poorly understood. In this study, we report on the characterization of two new ammonium transporters (AMT) in the perennial tree Pyrus betulaefolia. In silico analyses and yeast complementation assays revealed that both PbAMT1;3 and PbAMT1;5 can be classified in the AMT1 sub-family. The specific expression of PbAMT1;3 in roots and of PbAMT1;5 in leaves indicates that they have diverse functions in ammonium uptake or transport in P. betulaefolia. Their expression was strongly influenced by ammonium availability. In addition, the transcript level of PbAMT1;5 was significantly affected by the diurnal cycle and senescence hormones. They conferred the ability to uptake nitrogen to the yeast strain 31019b; however, the 15NH4 + uptake kinetics of PbAMT1;3 were different from those of PbAMT1;5. Indeed, PbAMT1;3 had a higher affinity for 15NH4 +, and pH changes were associated with this substrates’ transport in yeast. The present study provides basic gene features and transcriptional information for the two new members of the AMT1 sub-family in P. betulaefolia and will aid in decoding the precise roles of AMTs in P. betulaefolia physiology.

Keywords

Pyrus betulaefolia Ammonium transporter Gene characterization Expression pattern Functional complementation 

Abbreviations

ABA

Abscisic acid

AMT

Ammonium transporter

Arg

Arginine

MEGA

Molecular evolutionary genetics analysis

MeJ

Methyl jasmonate

OD600

Absorbance value at 600 nm

qPCR

Quantitative real-time PCR

TMs

Transmembrane domains

Notes

Acknowledgments

This study was supported by the National Natural Science Foundation of China (Grant Number: 31372051).

Supplementary material

10265_2016_799_MOESM1_ESM.pdf (319 kb)
Supplementary material 1 (PDF 320 kb)

References

  1. Becker D, Stanke R, Fendrik I, Frommer WB, Vanderleyden J, Kaiser WM, Hedrich R (2002) Expression of the NH4 +-transporter gene LEAMT1;2 is induced in tomato roots upon association with N2-fixing bacteria. Planta 215:424–429CrossRefPubMedGoogle Scholar
  2. Britto DT, Glass ADM, Kronzucker HJ, Siddiqi MY (2001) Cytosolic concentrations and transmembrane fluxes of NH4 +/NH3. An evaluation of recent proposals. Plant Physiol 125:523–526CrossRefPubMedPubMedCentralGoogle Scholar
  3. Couturier J, Montanini B, Martin F, Brun A, Blaudez D, Chalot M (2007) The expanded family of ammonium transporters in the perennial poplar plant. New Phytol 174:137–150CrossRefPubMedGoogle Scholar
  4. D’Apuzzo E, Rogato A, Simon-Rosin U, El Alaoui H, Barbulova A, Betti M, Dimou M, Katinakis P, Marquez A, Marini AM, Udvardi MK, Chiurazzi M (2004) Characterization of three functional high-affinity ammonium transporters in Lotus japonicus with differential transcriptional regulation spatial expression. Plant Physiol 134:1763–1774CrossRefPubMedPubMedCentralGoogle Scholar
  5. Gazzarrini S, Lejay L, Gojon A, Ninnemann O, Frommer WB, von Wirén N (1999) Three functional transporters for constitutive, diurnally regulated, and starvation-induced uptake of ammonium into Arabidopsis roots. Plant Cell 11:937–948CrossRefPubMedPubMedCentralGoogle Scholar
  6. Gu R, Duan F, An X, Zhang F, von Wirén N, Yuan L (2013) Characterization of AMT-mediated high-affinity ammonium uptake in roots of maize (Zea mays L.). Plant Cell Physiol 54:1515–1524CrossRefPubMedGoogle Scholar
  7. Güether M, Neuhaüser B, Balestrini R, Dynowski M, Ludewig U, Bonfante P (2009) A mycorrhizal-specific ammonium transporter from Lotus japonicus acquires nitrogen released by arbuscular mycorrhizal fungi. Plant Physiol 150:73–83CrossRefPubMedPubMedCentralGoogle Scholar
  8. Lauter FR, Ninnemann O, Bucher M, Riesmeier JW, Frommer WB (1996) Preferential expression of an ammonium transporter and of two putative nitrate transporters in root hairs of tomato. PANS 93:8139–8144CrossRefGoogle Scholar
  9. Li H, Cong Y, Lin J, Chang YH (2015) Molecular cloning and identification of an ammonium transporter gene from pear. Plant Cell, Tissue Organ Cult 120:441–451CrossRefGoogle Scholar
  10. Loqué D, von Wirén N (2004) Regulatory levels for the transport of ammonium in plant roots. J Exp Bot 55:1293–1305CrossRefPubMedGoogle Scholar
  11. Ludewig U (2006) Ion transport versus gas conduction: function of AMT/Rh-type proteins. Transfus Clin Biol 13:111–116CrossRefPubMedGoogle Scholar
  12. Ludewig U, von Wirén N, Frommer WB (2002) Uniport of NH4 + by the root hair plasma membrane ammonium transporter LeAMT1;1. J Biol Chem 277:13548–13555CrossRefPubMedGoogle Scholar
  13. Ludewig U, Wilken S, Wu BH, Jost W, Obrdlik P, El Bakkoury M, Marini AM, Andre B, Hamacher T, Boles E, Frommer WB, von Wirén N (2003) Homo- and hetero-oligomerization of ammonium transporter-1 NH4 + uniporters. J Biol Chem 278:45603–45610CrossRefPubMedGoogle Scholar
  14. Ludewig U, Neuhduser B, Dynowski M (2007) Molecular mechanisms of ammonium transport and accumulation in plants. FEBS Lett 581:2301–2308CrossRefPubMedGoogle Scholar
  15. Marini AM, SoussiBoudekou S, Vissers S, Andre B (1997) A family of ammonium transporters in Saccharomyces cerevisiae. Mol Cell Biol 17:4282–4293CrossRefPubMedPubMedCentralGoogle Scholar
  16. Mayer M, Ludewig U (2006) Role of AMT1;1 in NH4 + acquisition in Arabidopsis thaliana. Plant Biol 8:522–528CrossRefPubMedGoogle Scholar
  17. Mayer M, Dynowski M, Ludewig U (2006) Ammonium ion transport by the AMT/Rh homologue LeAMT1;1. Biochem J 396:431–437CrossRefPubMedPubMedCentralGoogle Scholar
  18. McDonald TR, Dietrich FS, Lutzoni F (2012) Multiple horizontal gene transfers of ammonium transporters/ammonia permeases from prokaryotes to eukaryotes: toward a new functional and evolutionary classification. Mol Biol Evol 29:51–60CrossRefPubMedGoogle Scholar
  19. Neuhäuser B, Dynowski M, Mayer M, Ludewig U (2007) Regulation of NH4 + transport by essential cross talk between AMT monomers through the carboxyl tails. Plant Physiol 143:1651–1659CrossRefPubMedPubMedCentralGoogle Scholar
  20. Ninnemann O, Jauniaux JC, Frommer WB (1994) Identification of a high-affinity NH4 + transporter from plants. EMBO J 13:3464–3471PubMedPubMedCentralGoogle Scholar
  21. Ortiz-Ramirez C, Mora SI, Trejo J, Pantoja O (2011) PvAMT1;1, a highly selective ammonium transporter that functions as H+/NH4 + symporter. J Biol Chem 286:31113–31122CrossRefPubMedPubMedCentralGoogle Scholar
  22. Salvemini F, Marini AM, Riccio A, Patriarca EJ, Chiurazzi M (2001) Functional characterization of an ammonium transporter gene from Lotus japonicus. Gene 270:237–243CrossRefPubMedGoogle Scholar
  23. Selle A, Willmann M, Grunze N, Gessler A, Weiss M, Nehls U (2005) The high-affinity poplar ammonium importer PttAMT1.2 and its role in ectomycorrhizal symbiosis. New Phytol 168:697–706CrossRefPubMedGoogle Scholar
  24. Shelden MC, Dong B, de Bruxelles GL, Trevaskis B, Whelan J, Ryan PR, Howitt SM, Udvardi MK (2001) Arabidopsis ammonium transporters, AtAMT1;1 and AtAMT1;2, have different biochemical properties and functional roles. Plant Soil 231:151–160CrossRefGoogle Scholar
  25. Sogaard R, Alsterfjord M, MacAulay N, Zeuthen T (2009) Ammonium ion transport by the AMT/Rh homolog TaAMT1;1 is stimulated by acidic pH. Pflug Arch Eur J Physiol 458:733–743CrossRefGoogle Scholar
  26. Sonoda Y, Ikeda A, Saiki S, von Wirén N, Yamaya T, Yamaguchi J (2003) Distinct expression and function of three ammonium transporter genes (OsAMT1;1-1;3) in rice. Plant Cell Physiol 44:726–734CrossRefPubMedGoogle Scholar
  27. Soupene E, Ramirez RM, Kustu S (2001) Evidence that fungal MEP proteins mediate diffusion of the uncharged species NH3 across the cytoplasmic membrane. Mol Cell Biol 21(17):5733–5741CrossRefPubMedPubMedCentralGoogle Scholar
  28. Straub D, Ludewig U, Neuhäuser B (2014) A nitrogen-dependent switch in the high affinity ammonium transport in Medicago truncatula. Plant Mol Biol 86:485–494CrossRefPubMedGoogle Scholar
  29. 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 28:2731–2739CrossRefPubMedPubMedCentralGoogle Scholar
  30. Thomas GH, Mullins JG, Merrick M (2000) Membrane topology of the Mep/Amt family of ammonium transporters. Mol Microbiol 37:331–344CrossRefPubMedGoogle Scholar
  31. van der Graaff E, Schwacke R, Schneider A, Desimone M, Flugge UI, Kunze R (2006) Transcription analysis of Arabidopsis membrane transporters and hormone pathways during developmental and induced leaf senescence. Plant Physiol 141:776–792CrossRefPubMedPubMedCentralGoogle Scholar
  32. von Wirén N, Lauter FR, Ninnemann O, Gillissen B, Walch-Liu P, Engels C, Jost W, Frommer WB (2000) Differential regulation of three functional ammonium transporter genes by nitrogen in root hairs and by light in leaves of tomato. Plant J 21:167–175CrossRefGoogle Scholar
  33. Yuan L, Loqué D, Kojima S, Rauch S, Ishiyama K, Inoue E, Takahashi H, von Wirén N (2007) The organization of high-affinity ammonium uptake in Arabidopsis roots depends on the spatial arrangement and biochemical properties of AMT1-type transporters. Plant Cell 19:2636–2652CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© The Botanical Society of Japan and Springer Japan 2016

Authors and Affiliations

  • Hui Li
    • 1
  • Jin-Long Han
    • 1
  • You-Hong Chang
    • 1
  • Jing Lin
    • 1
  • Qing-Song Yang
    • 1
  1. 1.Institute of Horticulture, Jiangsu Academy of Agricultural Sciences/Jiangsu Key Laboratory for Horticultural Crop Genetic ImprovementNanjingChina

Personalised recommendations