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HvALMT1 from barley is involved in the transport of organic anions.

Gruber BD, Ryan PR, Richardson AE, Tyerman SD, Ramesh S, Hebb DM, Howitt SM, Delhaize E - J. Exp. Bot. (2010)

Bottom Line: HvALMT1 is located on chromosome 2H which has not been associated with Al(3+) resistance in barley.Both malate uptake and efflux were confirmed in oocyte assays using [(14)C]malate as a radiotracer.It is suggested that HvALMT1 functions as an anion channel to facilitate organic anion transport in stomatal function and expanding cells.

View Article: PubMed Central - PubMed

Affiliation: CSIRO Plant Industry, GPO Box 1600, Canberra, ACT, 2601, Australia.

ABSTRACT
Members of the ALMT gene family contribute to the Al(3+) resistance of several plant species by facilitating malate efflux from root cells. The first member of this family to be cloned and characterized, TaALMT1, is responsible for most of the natural variation of Al(3+) resistance in wheat. The current study describes the isolation and characterization of HvALMT1, the barley gene with the greatest sequence similarity to TaALMT1. HvALMT1 is located on chromosome 2H which has not been associated with Al(3+) resistance in barley. The relatively low levels of HvALMT1 expression detected in root and shoot tissues were independent of external aluminium or phosphorus supply. Transgenic barley plants transformed with the HvALMT1 promoter fused to the green fluorescent protein (GFP) indicated that expression of HvALMT1 was relatively high in stomatal guard cells and in root tissues containing expanding cells. GFP fused to the C-terminus of the full HvALMT1 protein localized to the plasma membrane and motile vesicles within the cytoplasm. HvALMT1 conferred both inward and outward currents when expressed in Xenopus laevis oocytes that were bathed in a range of anions including malate. Both malate uptake and efflux were confirmed in oocyte assays using [(14)C]malate as a radiotracer. It is suggested that HvALMT1 functions as an anion channel to facilitate organic anion transport in stomatal function and expanding cells.

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Related in: MedlinePlus

Efflux of radioactively labelled malate from Xenopus oocytes expressing HvALMT1. Oocytes were pre-loaded with [14C]malate and the radioactivity of the bathing solution was measured. Oocytes were injected with either HvALMT1 cRNA or water as a control and incubated at external pH 7.5 or pH 4.5. Shown is the mean efflux of radioactive malate from the oocyte 2 min and 5 min after injection. Error bars show the SEM (n=5), and columns with different letters are significantly different (P <0.05; n=5).
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fig6: Efflux of radioactively labelled malate from Xenopus oocytes expressing HvALMT1. Oocytes were pre-loaded with [14C]malate and the radioactivity of the bathing solution was measured. Oocytes were injected with either HvALMT1 cRNA or water as a control and incubated at external pH 7.5 or pH 4.5. Shown is the mean efflux of radioactive malate from the oocyte 2 min and 5 min after injection. Error bars show the SEM (n=5), and columns with different letters are significantly different (P <0.05; n=5).

Mentions: Malate efflux from oocytes was also monitored after they were injected with [14C]malate. In these experiments the malate efflux after 2 min was ∼2.5-fold greater in oocytes expressing HvALMT1 (pHext 4.5) than in control oocytes injected with water (Fig. 6). This difference was not apparent 5 min after injection or at time points up to 30 min later (data not shown for later time points). Furthermore, the efflux was not significantly different from that of control oocytes when the external pH was 7.5 rather than 4.5, which is consistent with the electrophysiological data. Malate uptake by the oocytes was monitored by bathing the oocytes in [14C]malate and measuring the radioactivity taken up. Although the difference was smaller than observed for efflux, the rate of malate uptake was significantly greater in oocytes expressing HvALMT1 than in oocytes injected with H2O at pHext 4.5, but not at pHext 7.5 (Fig. 7).


HvALMT1 from barley is involved in the transport of organic anions.

Gruber BD, Ryan PR, Richardson AE, Tyerman SD, Ramesh S, Hebb DM, Howitt SM, Delhaize E - J. Exp. Bot. (2010)

Efflux of radioactively labelled malate from Xenopus oocytes expressing HvALMT1. Oocytes were pre-loaded with [14C]malate and the radioactivity of the bathing solution was measured. Oocytes were injected with either HvALMT1 cRNA or water as a control and incubated at external pH 7.5 or pH 4.5. Shown is the mean efflux of radioactive malate from the oocyte 2 min and 5 min after injection. Error bars show the SEM (n=5), and columns with different letters are significantly different (P <0.05; n=5).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2837267&req=5

fig6: Efflux of radioactively labelled malate from Xenopus oocytes expressing HvALMT1. Oocytes were pre-loaded with [14C]malate and the radioactivity of the bathing solution was measured. Oocytes were injected with either HvALMT1 cRNA or water as a control and incubated at external pH 7.5 or pH 4.5. Shown is the mean efflux of radioactive malate from the oocyte 2 min and 5 min after injection. Error bars show the SEM (n=5), and columns with different letters are significantly different (P <0.05; n=5).
Mentions: Malate efflux from oocytes was also monitored after they were injected with [14C]malate. In these experiments the malate efflux after 2 min was ∼2.5-fold greater in oocytes expressing HvALMT1 (pHext 4.5) than in control oocytes injected with water (Fig. 6). This difference was not apparent 5 min after injection or at time points up to 30 min later (data not shown for later time points). Furthermore, the efflux was not significantly different from that of control oocytes when the external pH was 7.5 rather than 4.5, which is consistent with the electrophysiological data. Malate uptake by the oocytes was monitored by bathing the oocytes in [14C]malate and measuring the radioactivity taken up. Although the difference was smaller than observed for efflux, the rate of malate uptake was significantly greater in oocytes expressing HvALMT1 than in oocytes injected with H2O at pHext 4.5, but not at pHext 7.5 (Fig. 7).

Bottom Line: HvALMT1 is located on chromosome 2H which has not been associated with Al(3+) resistance in barley.Both malate uptake and efflux were confirmed in oocyte assays using [(14)C]malate as a radiotracer.It is suggested that HvALMT1 functions as an anion channel to facilitate organic anion transport in stomatal function and expanding cells.

View Article: PubMed Central - PubMed

Affiliation: CSIRO Plant Industry, GPO Box 1600, Canberra, ACT, 2601, Australia.

ABSTRACT
Members of the ALMT gene family contribute to the Al(3+) resistance of several plant species by facilitating malate efflux from root cells. The first member of this family to be cloned and characterized, TaALMT1, is responsible for most of the natural variation of Al(3+) resistance in wheat. The current study describes the isolation and characterization of HvALMT1, the barley gene with the greatest sequence similarity to TaALMT1. HvALMT1 is located on chromosome 2H which has not been associated with Al(3+) resistance in barley. The relatively low levels of HvALMT1 expression detected in root and shoot tissues were independent of external aluminium or phosphorus supply. Transgenic barley plants transformed with the HvALMT1 promoter fused to the green fluorescent protein (GFP) indicated that expression of HvALMT1 was relatively high in stomatal guard cells and in root tissues containing expanding cells. GFP fused to the C-terminus of the full HvALMT1 protein localized to the plasma membrane and motile vesicles within the cytoplasm. HvALMT1 conferred both inward and outward currents when expressed in Xenopus laevis oocytes that were bathed in a range of anions including malate. Both malate uptake and efflux were confirmed in oocyte assays using [(14)C]malate as a radiotracer. It is suggested that HvALMT1 functions as an anion channel to facilitate organic anion transport in stomatal function and expanding cells.

Show MeSH
Related in: MedlinePlus