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Vacuolar-Iron-Transporter1-Like proteins mediate iron homeostasis in Arabidopsis.

Gollhofer J, Timofeev R, Lan P, Schmidt W, Buckhout TJ - PLoS ONE (2014)

Bottom Line: Isolated vacuoles from yeast expressing either of the VTL genes in the Δccc1 background had a three- to four-fold increase in Fe concentration compared to vacuoles isolated from the untransformed mutant.Seedling root growth of the Arabidopsis nramp3/nramp4 and vit1-1 mutants was decreased compared to the wild type when seedlings were grown under Fe deficiency.We conclude that the VTL proteins catalyze Fe transport into vacuoles and thus contribute to the regulation of Fe homeostasis in planta.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biology, Humboldt University Berlin, Berlin, Germany.

ABSTRACT
Iron deficiency is a nutritional problem in plants and reduces crop productivity, quality and yield. With the goal of improving the iron (Fe) storage properties of plants, we have investigated the function of three Arabidopsis proteins with homology to Vacuolar Iron Transporter1 (AtVIT1). Heterologous expression of Vacuolar Iron Transporter-Like1 (AtVTL1; At1g21140), AtVTL2 (At1g76800) or AtVTL5 (At3g25190) in the yeast vacuolar Fe transport mutant, Δccc1, restored growth in the presence of 4 mM Fe. Isolated vacuoles from yeast expressing either of the VTL genes in the Δccc1 background had a three- to four-fold increase in Fe concentration compared to vacuoles isolated from the untransformed mutant. Transiently expressed GFP-tagged AtVTL1 was localized exclusively and AtVTL2 was localized primarily to the vacuolar membrane of onion epidermis cells. Seedling root growth of the Arabidopsis nramp3/nramp4 and vit1-1 mutants was decreased compared to the wild type when seedlings were grown under Fe deficiency. When expressed under the 35S promoter in the nramp3/nramp4 or vit1-1 backgrounds, AtVTL1, AtVTL2 or AtVTL5 restored root growth in both mutants. The seed Fe concentration in the nramp3/nramp4 mutant overexpressing AtVTL1, AtVTL2 or AtVTL5 was between 50 and 60% higher than in non-transformed double mutants or wild-type plants. We conclude that the VTL proteins catalyze Fe transport into vacuoles and thus contribute to the regulation of Fe homeostasis in planta.

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Complementation of the Δccc1 by heterologous expression of the VTL genes.Δccc1 (vacuolar Fe2+/Mn2+ transporter) cells were transformed with each of the three VTL genes or the empty vector (pUE) under the control of the PGK promoter and grown on SD medium containing FeSO4 at the concentrations indicated for 24 or 48 h at 30°C. Cells were plated at the densities indicated in the figure.
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pone-0110468-g001: Complementation of the Δccc1 by heterologous expression of the VTL genes.Δccc1 (vacuolar Fe2+/Mn2+ transporter) cells were transformed with each of the three VTL genes or the empty vector (pUE) under the control of the PGK promoter and grown on SD medium containing FeSO4 at the concentrations indicated for 24 or 48 h at 30°C. Cells were plated at the densities indicated in the figure.

Mentions: As in Arabidopsis, the yeast vacuole is the major compartment for Fe storage but also protects the cell from Fe toxicity. The yeast gene CCC1 encodes a vacuolar Fe2+/Mn2+ transporter that catalyzes Fe2+ uptake into the vacuole. The Δccc1 mutant is hypersensitive to Fe toxicity, not growing in the presence of greater than ca. 3 mM Fe [25]. Because of the homology of the VTL family member to yeast Ccc1p and Arabidopsis AtVIT1 and because of the transcriptional repression of AtVTL1, AtVTL2 and AtVTL5 under conditions of Fe deficiency, the ability of the three AtVTL genes to complement the Δccc1 yeast mutant was investigated. When expressed in yeast, the VTL proteins were effective in complementing of the Δccc1 phenotype in the order AtVTL1> AtVTL2> AtVTL5 (Fig. 1). These results are consistent with an AtVTL-dependent decreased the cytosolic Fe in the Δccc1 mutant, presumably by transport of Fe into the vacuole.


Vacuolar-Iron-Transporter1-Like proteins mediate iron homeostasis in Arabidopsis.

Gollhofer J, Timofeev R, Lan P, Schmidt W, Buckhout TJ - PLoS ONE (2014)

Complementation of the Δccc1 by heterologous expression of the VTL genes.Δccc1 (vacuolar Fe2+/Mn2+ transporter) cells were transformed with each of the three VTL genes or the empty vector (pUE) under the control of the PGK promoter and grown on SD medium containing FeSO4 at the concentrations indicated for 24 or 48 h at 30°C. Cells were plated at the densities indicated in the figure.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215979&req=5

pone-0110468-g001: Complementation of the Δccc1 by heterologous expression of the VTL genes.Δccc1 (vacuolar Fe2+/Mn2+ transporter) cells were transformed with each of the three VTL genes or the empty vector (pUE) under the control of the PGK promoter and grown on SD medium containing FeSO4 at the concentrations indicated for 24 or 48 h at 30°C. Cells were plated at the densities indicated in the figure.
Mentions: As in Arabidopsis, the yeast vacuole is the major compartment for Fe storage but also protects the cell from Fe toxicity. The yeast gene CCC1 encodes a vacuolar Fe2+/Mn2+ transporter that catalyzes Fe2+ uptake into the vacuole. The Δccc1 mutant is hypersensitive to Fe toxicity, not growing in the presence of greater than ca. 3 mM Fe [25]. Because of the homology of the VTL family member to yeast Ccc1p and Arabidopsis AtVIT1 and because of the transcriptional repression of AtVTL1, AtVTL2 and AtVTL5 under conditions of Fe deficiency, the ability of the three AtVTL genes to complement the Δccc1 yeast mutant was investigated. When expressed in yeast, the VTL proteins were effective in complementing of the Δccc1 phenotype in the order AtVTL1> AtVTL2> AtVTL5 (Fig. 1). These results are consistent with an AtVTL-dependent decreased the cytosolic Fe in the Δccc1 mutant, presumably by transport of Fe into the vacuole.

Bottom Line: Isolated vacuoles from yeast expressing either of the VTL genes in the Δccc1 background had a three- to four-fold increase in Fe concentration compared to vacuoles isolated from the untransformed mutant.Seedling root growth of the Arabidopsis nramp3/nramp4 and vit1-1 mutants was decreased compared to the wild type when seedlings were grown under Fe deficiency.We conclude that the VTL proteins catalyze Fe transport into vacuoles and thus contribute to the regulation of Fe homeostasis in planta.

View Article: PubMed Central - PubMed

Affiliation: Institute of Biology, Humboldt University Berlin, Berlin, Germany.

ABSTRACT
Iron deficiency is a nutritional problem in plants and reduces crop productivity, quality and yield. With the goal of improving the iron (Fe) storage properties of plants, we have investigated the function of three Arabidopsis proteins with homology to Vacuolar Iron Transporter1 (AtVIT1). Heterologous expression of Vacuolar Iron Transporter-Like1 (AtVTL1; At1g21140), AtVTL2 (At1g76800) or AtVTL5 (At3g25190) in the yeast vacuolar Fe transport mutant, Δccc1, restored growth in the presence of 4 mM Fe. Isolated vacuoles from yeast expressing either of the VTL genes in the Δccc1 background had a three- to four-fold increase in Fe concentration compared to vacuoles isolated from the untransformed mutant. Transiently expressed GFP-tagged AtVTL1 was localized exclusively and AtVTL2 was localized primarily to the vacuolar membrane of onion epidermis cells. Seedling root growth of the Arabidopsis nramp3/nramp4 and vit1-1 mutants was decreased compared to the wild type when seedlings were grown under Fe deficiency. When expressed under the 35S promoter in the nramp3/nramp4 or vit1-1 backgrounds, AtVTL1, AtVTL2 or AtVTL5 restored root growth in both mutants. The seed Fe concentration in the nramp3/nramp4 mutant overexpressing AtVTL1, AtVTL2 or AtVTL5 was between 50 and 60% higher than in non-transformed double mutants or wild-type plants. We conclude that the VTL proteins catalyze Fe transport into vacuoles and thus contribute to the regulation of Fe homeostasis in planta.

Show MeSH
Related in: MedlinePlus