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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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Transient 35S:GFP::VTL1 and 35S:GFP::VTL2 expression in onion epidermis cells.The GFP-VTL1 fluorescent signal co-localized with the vacuolar marker, vac-rk CD3-975, in turgescent (A) and plasmolyzed (B) cells. The GFP-VTL2 signal was also observed on the plasma membrane and cytoplasma (B).
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pone-0110468-g003: Transient 35S:GFP::VTL1 and 35S:GFP::VTL2 expression in onion epidermis cells.The GFP-VTL1 fluorescent signal co-localized with the vacuolar marker, vac-rk CD3-975, in turgescent (A) and plasmolyzed (B) cells. The GFP-VTL2 signal was also observed on the plasma membrane and cytoplasma (B).

Mentions: The localization of AtVTL1 and AtVTL2 was investigated in onion epidermis cells. Attempts to localize AtVTL5 in onion or tobacco leaf cells have not been successful. Onion cells were transformed by particle bombardment with 35S:GFP::VTL1 or 35S:GFP::VTL2 reporter gene constructs. In the case of AtVTL1, the GFP fluorescence co-localized with the vacuolar marker vac-rk CD3-975 (Fig. 3A, [27]). When cells were plasmolyzed in 0.8 M mannitol, the GFP signal remained associated with the vacuole (Fig. 3B). The localization of AtVTL2 in epidermis cells was predominately associated with the vacuolar membrane (Fig. 3A); however, GFP fluorescence was also found to be associated with the plasma membrane and the cytoplasm upon plasmolysis (Fig. 3B). The localization of the two AtVTL proteins on the onion vacuolar membrane was consistent with the complementation results in Saccharomyces and with the increased Fe concentration in vacuoles isolated from VTL-expressing yeast cells.


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

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

Transient 35S:GFP::VTL1 and 35S:GFP::VTL2 expression in onion epidermis cells.The GFP-VTL1 fluorescent signal co-localized with the vacuolar marker, vac-rk CD3-975, in turgescent (A) and plasmolyzed (B) cells. The GFP-VTL2 signal was also observed on the plasma membrane and cytoplasma (B).
© Copyright Policy
Related In: Results  -  Collection

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

pone-0110468-g003: Transient 35S:GFP::VTL1 and 35S:GFP::VTL2 expression in onion epidermis cells.The GFP-VTL1 fluorescent signal co-localized with the vacuolar marker, vac-rk CD3-975, in turgescent (A) and plasmolyzed (B) cells. The GFP-VTL2 signal was also observed on the plasma membrane and cytoplasma (B).
Mentions: The localization of AtVTL1 and AtVTL2 was investigated in onion epidermis cells. Attempts to localize AtVTL5 in onion or tobacco leaf cells have not been successful. Onion cells were transformed by particle bombardment with 35S:GFP::VTL1 or 35S:GFP::VTL2 reporter gene constructs. In the case of AtVTL1, the GFP fluorescence co-localized with the vacuolar marker vac-rk CD3-975 (Fig. 3A, [27]). When cells were plasmolyzed in 0.8 M mannitol, the GFP signal remained associated with the vacuole (Fig. 3B). The localization of AtVTL2 in epidermis cells was predominately associated with the vacuolar membrane (Fig. 3A); however, GFP fluorescence was also found to be associated with the plasma membrane and the cytoplasm upon plasmolysis (Fig. 3B). The localization of the two AtVTL proteins on the onion vacuolar membrane was consistent with the complementation results in Saccharomyces and with the increased Fe concentration in vacuoles isolated from VTL-expressing yeast cells.

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