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Arabidopsis thaliana POLYOL/MONOSACCHARIDE TRANSPORTERS 1 and 2: fructose and xylitol/H+ symporters in pollen and young xylem cells.

Klepek YS, Volke M, Konrad KR, Wippel K, Hoth S, Hedrich R, Sauer N - J. Exp. Bot. (2009)

Bottom Line: Analyses of reporter genes performed with AtPMT1 or AtPMT2 promoter sequences showed expression in mature (AtPMT2) or germinating (AtPMT1) pollen grains, as well as in growing pollen tubes, hydathodes, and young xylem cells (both genes).The expression was confirmed with an anti-AtPMT1/AtPMT2 antiserum (alphaAtPMT1/2) raised against peptides conserved in AtPMT1 and AtPMT2.The physiological roles of the proteins are discussed and related to plant cell wall modifications.

View Article: PubMed Central - PubMed

Affiliation: Molekulare Pflanzenphysiologie, Universität Erlangen-Nürnberg, Staudtstrasse 5, Erlangen, Germany.

ABSTRACT
The genome of Arabidopsis thaliana contains six genes, AtPMT1 to AtPMT6 (Arabidopsis thaliana POLYOL/MONOSACCHARIDE TRANSPORTER 1-6), which form a distinct subfamily within the large family of more than 50 monosaccharide transporter-like (MST-like) genes. So far, only AtPMT5 [formerly named AtPLT5 (At3g18830)] has been characterized and was shown to be a plasma membrane-localized H(+)-symporter with broad substrate specificity. The characterization of AtPMT1 (At2g16120) and AtPMT2 (At2g16130), two other, almost identical, members of this transporter subfamily, are presented here. Expression of the AtPMT1 and AtPMT2 cDNAs in baker's yeast (Saccharomyces cerevisiae) revealed that these proteins catalyse the energy-dependent, high-capacity transport of fructose and xylitol, and the transport of several other compounds with lower rates. Expression of their cRNAs in Xenopus laevis oocytes showed that both proteins are voltage-dependent and catalyse the symport of their substrates with protons. Fusions of AtPMT1 or AtPMT2 with the green fluorescent protein (GFP) localized to Arabidopsis plasma membranes. Analyses of reporter genes performed with AtPMT1 or AtPMT2 promoter sequences showed expression in mature (AtPMT2) or germinating (AtPMT1) pollen grains, as well as in growing pollen tubes, hydathodes, and young xylem cells (both genes). The expression was confirmed with an anti-AtPMT1/AtPMT2 antiserum (alphaAtPMT1/2) raised against peptides conserved in AtPMT1 and AtPMT2. The physiological roles of the proteins are discussed and related to plant cell wall modifications.

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Reporter gene analyses of pAtPMT2/GFP and pAtPMT2/GUS plants. (A) Inflorescence of a pAtPMT2/GUS plant with strong GUS staining in the mature anthers. (B) Higher magnification of an anther with very strong GUS staining in fully developed pollen grains (arrows) and in germinated and ungerminated pollen on agar medium (insert). Staining of cells in the anther surface results from the diffusion of excess stain out of the pollen grains and can even reach the sepals and petals of stained flowers (see A). (C) GFP-fluorescence (epifluorescence) in pollen grains on an opened anther from a pAtPMT2/GFP plants. No fluorescence is seen in WT anthers (insert). (D) Strong GUS staining in source leaf hydathodes and very weak GUS staining in minor veins (arrows). (E) Cross-section of a flower stalk with GUS staining in young xylem cells (ca, cambium; ph, phloem; xy, xylem). A bar indicates the region where the single-celled row of cambial cells is located. The cambial cells themselves cannot be identified. Bars are 2 mm (A, D), 200 μm (B, insert of C), 20 μm (insert of B), 100 μm (C), 25 μm (E).
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fig7: Reporter gene analyses of pAtPMT2/GFP and pAtPMT2/GUS plants. (A) Inflorescence of a pAtPMT2/GUS plant with strong GUS staining in the mature anthers. (B) Higher magnification of an anther with very strong GUS staining in fully developed pollen grains (arrows) and in germinated and ungerminated pollen on agar medium (insert). Staining of cells in the anther surface results from the diffusion of excess stain out of the pollen grains and can even reach the sepals and petals of stained flowers (see A). (C) GFP-fluorescence (epifluorescence) in pollen grains on an opened anther from a pAtPMT2/GFP plants. No fluorescence is seen in WT anthers (insert). (D) Strong GUS staining in source leaf hydathodes and very weak GUS staining in minor veins (arrows). (E) Cross-section of a flower stalk with GUS staining in young xylem cells (ca, cambium; ph, phloem; xy, xylem). A bar indicates the region where the single-celled row of cambial cells is located. The cambial cells themselves cannot be identified. Bars are 2 mm (A, D), 200 μm (B, insert of C), 20 μm (insert of B), 100 μm (C), 25 μm (E).

Mentions: Analysis of 30 pAtPMT2/GUS plants confirmed the already described GFP expression in pollen (Fig. 7A, B). In contrast to the GUS staining observed in pAtPMT1/GUS pollen (Fig. 6B, C), however, this staining (i) was much stronger and (ii) was detected in pollen grains of still closed anthers (Fig. 7B). Moreover, our analyses also revealed GUS activity in hydathodes and (weakly) in minor veins of several plants (Fig. 7D). Finally, one pAtPMT2/GUS plant showed staining in the vasculature of the flower stalks. The latter signal resulted from low GUS activity in the cambium and from stronger GUS activity in young xylem cells (Fig. 7E).


Arabidopsis thaliana POLYOL/MONOSACCHARIDE TRANSPORTERS 1 and 2: fructose and xylitol/H+ symporters in pollen and young xylem cells.

Klepek YS, Volke M, Konrad KR, Wippel K, Hoth S, Hedrich R, Sauer N - J. Exp. Bot. (2009)

Reporter gene analyses of pAtPMT2/GFP and pAtPMT2/GUS plants. (A) Inflorescence of a pAtPMT2/GUS plant with strong GUS staining in the mature anthers. (B) Higher magnification of an anther with very strong GUS staining in fully developed pollen grains (arrows) and in germinated and ungerminated pollen on agar medium (insert). Staining of cells in the anther surface results from the diffusion of excess stain out of the pollen grains and can even reach the sepals and petals of stained flowers (see A). (C) GFP-fluorescence (epifluorescence) in pollen grains on an opened anther from a pAtPMT2/GFP plants. No fluorescence is seen in WT anthers (insert). (D) Strong GUS staining in source leaf hydathodes and very weak GUS staining in minor veins (arrows). (E) Cross-section of a flower stalk with GUS staining in young xylem cells (ca, cambium; ph, phloem; xy, xylem). A bar indicates the region where the single-celled row of cambial cells is located. The cambial cells themselves cannot be identified. Bars are 2 mm (A, D), 200 μm (B, insert of C), 20 μm (insert of B), 100 μm (C), 25 μm (E).
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2803217&req=5

fig7: Reporter gene analyses of pAtPMT2/GFP and pAtPMT2/GUS plants. (A) Inflorescence of a pAtPMT2/GUS plant with strong GUS staining in the mature anthers. (B) Higher magnification of an anther with very strong GUS staining in fully developed pollen grains (arrows) and in germinated and ungerminated pollen on agar medium (insert). Staining of cells in the anther surface results from the diffusion of excess stain out of the pollen grains and can even reach the sepals and petals of stained flowers (see A). (C) GFP-fluorescence (epifluorescence) in pollen grains on an opened anther from a pAtPMT2/GFP plants. No fluorescence is seen in WT anthers (insert). (D) Strong GUS staining in source leaf hydathodes and very weak GUS staining in minor veins (arrows). (E) Cross-section of a flower stalk with GUS staining in young xylem cells (ca, cambium; ph, phloem; xy, xylem). A bar indicates the region where the single-celled row of cambial cells is located. The cambial cells themselves cannot be identified. Bars are 2 mm (A, D), 200 μm (B, insert of C), 20 μm (insert of B), 100 μm (C), 25 μm (E).
Mentions: Analysis of 30 pAtPMT2/GUS plants confirmed the already described GFP expression in pollen (Fig. 7A, B). In contrast to the GUS staining observed in pAtPMT1/GUS pollen (Fig. 6B, C), however, this staining (i) was much stronger and (ii) was detected in pollen grains of still closed anthers (Fig. 7B). Moreover, our analyses also revealed GUS activity in hydathodes and (weakly) in minor veins of several plants (Fig. 7D). Finally, one pAtPMT2/GUS plant showed staining in the vasculature of the flower stalks. The latter signal resulted from low GUS activity in the cambium and from stronger GUS activity in young xylem cells (Fig. 7E).

Bottom Line: Analyses of reporter genes performed with AtPMT1 or AtPMT2 promoter sequences showed expression in mature (AtPMT2) or germinating (AtPMT1) pollen grains, as well as in growing pollen tubes, hydathodes, and young xylem cells (both genes).The expression was confirmed with an anti-AtPMT1/AtPMT2 antiserum (alphaAtPMT1/2) raised against peptides conserved in AtPMT1 and AtPMT2.The physiological roles of the proteins are discussed and related to plant cell wall modifications.

View Article: PubMed Central - PubMed

Affiliation: Molekulare Pflanzenphysiologie, Universität Erlangen-Nürnberg, Staudtstrasse 5, Erlangen, Germany.

ABSTRACT
The genome of Arabidopsis thaliana contains six genes, AtPMT1 to AtPMT6 (Arabidopsis thaliana POLYOL/MONOSACCHARIDE TRANSPORTER 1-6), which form a distinct subfamily within the large family of more than 50 monosaccharide transporter-like (MST-like) genes. So far, only AtPMT5 [formerly named AtPLT5 (At3g18830)] has been characterized and was shown to be a plasma membrane-localized H(+)-symporter with broad substrate specificity. The characterization of AtPMT1 (At2g16120) and AtPMT2 (At2g16130), two other, almost identical, members of this transporter subfamily, are presented here. Expression of the AtPMT1 and AtPMT2 cDNAs in baker's yeast (Saccharomyces cerevisiae) revealed that these proteins catalyse the energy-dependent, high-capacity transport of fructose and xylitol, and the transport of several other compounds with lower rates. Expression of their cRNAs in Xenopus laevis oocytes showed that both proteins are voltage-dependent and catalyse the symport of their substrates with protons. Fusions of AtPMT1 or AtPMT2 with the green fluorescent protein (GFP) localized to Arabidopsis plasma membranes. Analyses of reporter genes performed with AtPMT1 or AtPMT2 promoter sequences showed expression in mature (AtPMT2) or germinating (AtPMT1) pollen grains, as well as in growing pollen tubes, hydathodes, and young xylem cells (both genes). The expression was confirmed with an anti-AtPMT1/AtPMT2 antiserum (alphaAtPMT1/2) raised against peptides conserved in AtPMT1 and AtPMT2. The physiological roles of the proteins are discussed and related to plant cell wall modifications.

Show MeSH
Related in: MedlinePlus