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Does a voltage-sensitive outer envelope transport mechanism contributes to the chloroplast iron uptake?


Main conclusion

Based on the effects of inorganic salts on chloroplast Fe uptake, the presence of a voltage-dependent step is proposed to play a role in Fe uptake through the outer envelope.

Although iron (Fe) plays a crucial role in chloroplast physiology, only few pieces of information are available on the mechanisms of chloroplast Fe acquisition. Here, the effect of inorganic salts on the Fe uptake of intact chloroplasts was tested, assessing Fe and transition metal uptake using bathophenantroline-based spectrophotometric detection and plasma emission-coupled mass spectrometry, respectively. The microenvironment of Fe was studied by Mössbauer spectroscopy. Transition metal cations (Cd2+, Zn2+, and Mn2+) enhanced, whereas oxoanions (NO3 , SO4 2−, and BO3 3−) reduced the chloroplast Fe uptake. The effect was insensitive to diuron (DCMU), an inhibitor of chloroplast inner envelope-associated Fe uptake. The inorganic salts affected neither Fe forms in the uptake assay buffer nor those incorporated into the chloroplasts. The significantly lower Zn and Mn uptake compared to that of Fe indicates that different mechanisms/transporters are involved in their acquisition. The enhancing effect of transition metals on chloroplast Fe uptake is likely related to outer envelope-associated processes, since divalent metal cations are known to inhibit Fe2+ transport across the inner envelope. Thus, a voltage-dependent step is proposed to play a role in Fe uptake through the chloroplast outer envelope on the basis of the contrasting effects of transition metal cations and oxoaninons.

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Light Harvesting Complex II apoprotein


Carbonyl cyanide m-chlorophenyl-hydrazone




Transmembrane electrochemical potential


Inner envelope


Outer envelope


Rubisco large subunit


  1. Abadía J, Vázquez S, Rellán-Álvarez R, El-Jendoubi H, Abadía A, Álvarez-Fernández A, López-Millán A-F (2011) Towards a knowledge-based correction of iron chlorosis. Plant Physiol Biochem 49:471–482

  2. Abdel-Ghany SE, Ye H, Garifullina GF, Zhang L, Pilon-Smits EAH, Pilon M (2005) Iron-sulfur cluster biogenesis in chloroplasts. Involvement of the scaffold protein CpIscA1. Plant Physiol 138:161–172

  3. Álvarez-Fernández A, Díaz-Benito P, Abadía A, López-Millán AF, Abadía J (2014) Metal species involved in long distance metal transport in plants. Front Plant Sci 5:105

  4. Andaluz S, López-Millán A-F, De las Rivas J, Aro E-M, Abadía J, Abadía A (2006) Proteomic profiles of thylakoid membranes and changes in response to iron deficiency. Photosynth Res 89:141–155

  5. Basa B, Lattanzio G, Solti Á, Tóth B, Abadía J, Fodor F, Sárvári É (2014) Changes induced by cadmium stress and iron deficiency in the composition and organization of thylakoid complexes in sugar beet (Beta vulgaris L.). Environ Exp Bot 101:1–11

  6. Bölter B, Soll J (2001) Ion channels in the outer membranes of chloroplasts and mitochondria: open doors or regulated gates? EMBO J 20:935–940

  7. Braun V (2003) Iron uptake by Escherichia coli. Front Biosci 8:1409–1421

  8. Braun V (2014) Energy-coupled transport across the outer membrane of Gram-negative bacteria. In: Remaut H, Frozens R (eds) Bacterial membranes: Structural and molecular biology. Caister Academic Press, Norfolk, pp 249–282

  9. Braun V, Hantke K (2011) Recent insights into iron import by bacteria. Curr Opin Chem Biol 15:328–334

  10. Braun V, Herrmann C (2007) Docking of the periplasmic FecB binding protein to the FecCD transmembrane proteins in the ferric citrate transport system of Escherichia coli. J Bacteriol 189:6913–6918

  11. Breuers FKH, Bräutigam A, Weber APM (2011) The plastid outer envelope–a highly dynamic interface between plastid and cytoplasm. Front Plant Scie 2:97

  12. Bughio N, Takahashi M, Yoshimuri E, Nishizawa N-K, Mori S (1997) Light-dependent iron transport into isolated barley chloroplasts. Plant Cell Physiol 38:101–105

  13. Castagna A, Donnini S, Ranieri A (2009) Adaptation to iron-deficiency requires remodelling of plant metabolism: an insight in chloroplast biochemistry and functionality. In: Ashraf M, Ozturk, H-u-R Athar M (eds) Salinity and water stress. Springer Verlag, Dordrecht, p 205–212

  14. Clausen C, Ilkavets I, Thomson R, Philippar K, Vojta A, Möhlmann T, Neuhaus E, Fulgosi H, Soll J (2004) Intracellular localization of VDAC proteins in plants. Planta 220:30–37

  15. Duy D, Soll J, Philippar K (2007a) Solute channels of the outer membrane: from bacteria to chloroplasts. Biol Chem 388:879–889

  16. Duy D, Wanner G, Meda AR, von Wirén N, Soll J, Philippar K (2007b) PIC1, an ancient permease in Arabidopsis chloroplasts, mediates iron transport. Plant Cell 19:986–1006

  17. Duy D, Stübe R, Wanner G, Philippar K (2011) The chloroplast permease PIC1 regulates plant growth and development by directing homeostasis and transport of iron. Plant Physiol 155:1709–1722

  18. Finazzi G, Petroutsos D, Tomizioli M, Flori S, Sautron E, Villanova V, Rolland N, Seigneurin-Berny D (2014) Ions channels/transporters and chloroplast regulation. Cell Calcium. doi:10.1016/j.ceca.2014.10.002

  19. Fodor F (2002) Physiological responses of vascular plants to heavy metals. In: Prasad MNV, Strzalka K (eds) Physiology and biochemistry of metal toxicity and tolerance in plants. Kluwer Academic Publisher, The Netherlands, pp 149–177

  20. Greenwood NN, Gibb TC (1971) Mössbauer spectroscopy. Chapman and Hall, London

  21. Gutierrez-Carbonell E, Takahashi D, Lattanzio G, Rodríguez-Celma J, Kehr J, Soll J, Philippar K, Uemura M, Abadía J, López-Millán AF (2014) The distinct functional roles of the inner and outer chloroplast envelope of pea (Pisum sativum) as revealed by proteomic approaches. J Prot Res 13:2941–2953

  22. Homblé F, Krammer E-M, Prévost M (2012) Plant VDAC: facts and speculations. Biochim Biophys Acta 1818:1486–1501

  23. Inoue K (2007) The chloroplast outer envelope membrane: The edge of light and excitement. J Integrat Plant Biol 49:1100–1111

  24. Inoue K (2011) Emerging roles of the chloroplast outer envelope membrane. Trend Plant Sci 16:1360–1385

  25. Jeong J, Cohu C, Kerkeb L, Pilon M, Connolly EL, Guerinot ML (2008) Chloroplast Fe(III) chelate reductase activity is essential for seedling viability under iron limiting conditions. Proc Natl Acad Sci USA 105:10619–10624

  26. Kim YY, Choi H, Segami S, Cho HT, Martinoia E, Maeshima M, Lee Y (2009) AtHMA1 contributes to the detoxification of excess Zn(II) in Arabidopsis. Plant J 58:737–753

  27. Klencsár Z, Kuzmann E, Vértes A (1996) User-friendly software for Mössbauer spectrum analysis. J Radioanal Nucl Chem 210:105–118

  28. Kobayashi T, Nishizawa NK (2012) Iron uptake, translocation, and regulation in higher plants. Annu Rev Plant Biol 63:131–152

  29. Krämer U, Talke IN, Hanikenne M (2007) Transition metal transport. FEBS Lett 581:2263–2272

  30. Lang EG, Mueller SJ, Hoernstein SN, Porankiewicz-Asplund J, Vervliet-Scheebaum M, Reski R (2011) Simultaneous isolation of pure and intact chloroplasts and mitochondria from moss as the basis for subcellular proteomics. Plant Cell Rep 30:205–215

  31. Latifi A, Jeanjean R, Lemeille S, Havaux M, Zhang C-C (2005) Iron starvation leads to oxidative stress in Anabaena sp. strain PCC 7120. J Bacteriol 187:6596–6598

  32. López-Millán AF, Duy D, Philippar K (2016) Chloroplast iron transport proteins–function and impact on plant physiology. Front Plant Sci 7:178

  33. Marshall B, Stintzi A, Gilmour G, Meyer J-M, Poole K (2009) Citrate-mediated iron uptake in Pseudomonas aeruginosa: involvement of the citrate-inducible FecA receptor and the FeoB ferrous iron transporter. Microbiol 155:305–315

  34. Moreau S, Day DA, Puppo A (1998) Ferrous iron is transported across the peribacteroid membrane of soybean nodules. Planta 207:83–87

  35. Morrissey J, Guerinot ML (2009) Iron uptake and transport in plants: the good, the bad, and the ionome. Chem Rev 109:4553–4567

  36. Nikolić M, Römheld V (2007) The dynamics of iron in the leaf apoplast significance for the iron nutrition of plants. In: Sattelmacher B, Horst WJ (eds) The apoplast of higher plants: Compartment of storage, transport and reactions. The significance of the apoplast for the mineral nutrition of higher plants. Springer Verlag, Dordrecht, pp 353–371

  37. Nouet C, Motte P, Hanikenne M (2011) Chloroplastic and mitochondrial metal homeostasis. Trend Plant Sci 16:1360–1385

  38. Palmer CM, Guerinot ML (2009) Facing the challenges of Cu, Fe and Zn homeostasis in plants. Nat Chem Biol 5:333–340

  39. Perfus-Barbeoch L, Leonhardt N, Vavasseur A, Forestier C (2002) Heavy metal toxicity: cadmium permeates through calcium channels and disturbs the plant water status. Plant J 32:539–548

  40. Pilon M, Cohu CM, Ravet K, Abdel-Ghany SE, Gaymard F (2009) Essential transition metal homeostasis in plants. Curr Opin Plant Biol 12:347–357

  41. Pottosin I, Dobrovinskaya O (2015) Ion channels in native chloroplast membranes: challenges and potential for direct patch-clamp studies. Front Physiol 6:396

  42. Rellán-Alvarez R, Giner-Martínez-Sierra J, Orduna J, Orera I, Rodríguez-Castrillón JA, García-Alonso JI, Abadía J, Alvarez-Fernández A (2010) Identification of a tri-iron(III), tri-citrate complex in the xylem sap of iron-deficient tomato resupplied with iron: new insights into plant iron long-distance transport. Plant Cell Physiol 51:91–102

  43. Reumann S, Keegstra K (1999) The endosymbiotic origin of the protein import machinery of chloroplastic envelope membranes. Trend Plant Sci 4:1360–1385

  44. Rodríguez-Serrano M, Romero-Puertas MC, Pazmiňo DM, Testillano PS, Risueňo MC, del Río LA, Sandalio LM (2009) Cellular response of pea plants to cadmium toxicity: cross talk between reactive oxygen species, nitric oxide, and calcium. Plant Physiol 150:229–243

  45. Röhl T, Motzkus M, Soll J (1999) The outer envelope protein OEP24 from pea chloroplasts can functionally replace the mitochondrial VDAC in yeast. FEBS Lett 460:491–494

  46. Shcolnick S, Keren N (2006) Metal homeostasis in cyanobacteria and chloroplasts. Balancing benefits and risks to the photosynthetic apparatus. Plant Physiol 141:805–810

  47. Shingles R, McCarty RE (1994) Direct measurement of ATP-dependent proton concentration changes and characterization of a K+-stimulated ATPase in pea chloroplast inner envelope vesicles. Plant Physiol 106:731–737

  48. Shingles R, North M, McCarty RE (2002) Ferrous ion transport across chloroplast inner envelope membranes. Plant Physiol 128:1022–1030

  49. Smith GF, McCurdy WH, Diehl H (1952) The colorimetric determination of iron in raw and treated municipal water supplies by use of 4:7-diphenyl-1:10-phenanthroline. Analyst 77:418–422

  50. Soll J, Bölter B, Wagner R, Hinnah SC (2000) The chloroplast outer envelope: a molecular sieve? Trends Plant Sci 5:137–138

  51. Solti Á, Kovács K, Basa B, Vértes A, Sárvári É, Fodor F (2012) Iron uptake of chloroplasts: kinetics, mechanism and incorporation. Plant Physiol Biochem 52:91–97

  52. Solti Á, Müller B, Czech V, Sárvári É, Fodor F (2014) Functional characterization of the chloroplast ferric chelate oxidoreductase enzyme. New Phytol 202:920–928

  53. Teng Y-S, Su Y-A, Chen L-J, Lee YJ, Hwang I, Li H-M (2006) Tic21 is an essential translocon component for protein translocation across the chloroplast inner envelope membrane. Plant Cell 18:2247–2257

  54. Terry N, Abadia J (1986) Function of iron in chloroplasts. J Plant Nutr 9:609–646

  55. Tewari RK, Kumar P, Neetu Sharma PN (2005) Signs of oxidative stress in the chlorotic leaves of iron starved plants. Plant Sci 169:1037–1045

  56. Timperio AM, D’Amici GM, Barta C, Loret F, Zolla L (2007) Proteomics, pigment composition, and organization of thylakoid membranes in iron-deficient spinach leaves. J Exp Bot 58:3695–3710

  57. Wang P, Kinraide TB, Zhou D, Kopittke PM, Peijnenburg WJGM (2011) Plasma membrane surface potential: dual effects upon ion uptake and toxicity. Plant Physiol 155:808–820

  58. Weber G, von Wirén N, Hayen H (2007) Investigation of ascorbate-mediated iron release from ferric phytosiderophores in the presence of nicotianamine. Biometals 21:503–513

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This work was supported by the Grants financed by the National Office for Research, Development, and Innovation, Hungary (OTKA-NKFIH PD-112047 and PD-111979), the Spanish Ministry of Economy and Competitivity (Project AGL2013–42175-R, co-financed with FEDER) and the Aragón Government (Group A03) to J.A. Á.S. was also supported by the Bolyai János Research Scholarship of the Hungarian Academy of Sciences (BO/00207/15/4).

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Correspondence to Ádám Solti.

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Solti, Á., Kovács, K., Müller, B. et al. Does a voltage-sensitive outer envelope transport mechanism contributes to the chloroplast iron uptake?. Planta 244, 1303–1313 (2016). https://doi.org/10.1007/s00425-016-2586-3

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  • Chloroplast
  • Envelope membrane
  • Iron metabolism
  • Mössbauer spectrosopy
  • Voltage-dependent transport