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Nitrogen affects cluster root formation and expression of putative peptide transporters.

Paungfoo-Lonhienne C, Schenk PM, Lonhienne TG, Brackin R, Meier S, Rentsch D, Schmidt S - J. Exp. Bot. (2009)

Bottom Line: The results provide further evidence that subgroup II of the NRT1/PTR family contains functional di- and tripeptide transporters.Grown in heathland or hydroponic culture with limiting N supply or starved of nutrients, HaPTR genes had the highest expression in cluster roots and non-cluster roots, and leaf expression increased upon re-supply of ON.It is concluded that formation of cluster roots and expression of PTR are regulated in response to N supply.

View Article: PubMed Central - PubMed

Affiliation: School of Integrative Biology, University of Queensland, QLD, 4072, Australia. chanyarat@uq.edu.au

ABSTRACT
Non-mycorrhizal Hakea actites (Proteaceae) grows in heathland where organic nitrogen (ON) dominates the soil nitrogen (N) pool. Hakea actites uses ON for growth, but the role of cluster roots in ON acquisition is unknown. The aim of the present study was to ascertain how N form and concentration affect cluster root formation and expression of peptide transporters. Hydroponically grown plants produced most biomass with low molecular weight ON>inorganic N>high molecular weight ON, while cluster roots were formed in the order no-N>ON>inorganic N. Intact dipeptide was transported into roots and metabolized, suggesting a role for the peptide transporter (PTR) for uptake and transport of peptides. HaPTR4, a member of subgroup II of the NRT1/PTR transporter family, which contains most characterized di- and tripeptide transporters in plants, facilitated transport of di- and tripeptides when expressed in yeast. No transport activity was demonstrated for HaPTR5 and HaPTR12, most similar to less well characterized transporters in subgroup III. The results provide further evidence that subgroup II of the NRT1/PTR family contains functional di- and tripeptide transporters. Green fluorescent protein fusion proteins of HaPTR4 and HaPTR12 localized to tonoplast, and plasma- and endomembranes, respectively, while HaPTR5 localized to vesicles of unknown identity. Grown in heathland or hydroponic culture with limiting N supply or starved of nutrients, HaPTR genes had the highest expression in cluster roots and non-cluster roots, and leaf expression increased upon re-supply of ON. It is concluded that formation of cluster roots and expression of PTR are regulated in response to N supply.

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Localization of fusion proteins of HaPTR with GFP in tobacco (Nicotiana tabacum) protoplasts. Confocal laser scanning microscope pictures (left) and corresponding bright field images (right) of tobacco protoplasts transiently expressing fusion proteins of HaPTRs with GFP or free GFP. Free GFP (A), ProT2–GFP (B), HaPTR4–GFP (C), HaPTR5–GFP (D), GFP–HaPTR12 (E), and HaPTR12–GFP (F). Merged images show GFP fluorescence (green) and chlorophyll fluorescence (red). Protoplast diameter ∼40 μm.
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fig4: Localization of fusion proteins of HaPTR with GFP in tobacco (Nicotiana tabacum) protoplasts. Confocal laser scanning microscope pictures (left) and corresponding bright field images (right) of tobacco protoplasts transiently expressing fusion proteins of HaPTRs with GFP or free GFP. Free GFP (A), ProT2–GFP (B), HaPTR4–GFP (C), HaPTR5–GFP (D), GFP–HaPTR12 (E), and HaPTR12–GFP (F). Merged images show GFP fluorescence (green) and chlorophyll fluorescence (red). Protoplast diameter ∼40 μm.

Mentions: To demonstrate intracellular localization, GFP fusion proteins were transiently expressed in tobacco protoplasts. Free GFP showed characteristic fluorescence in the cytosol (Fig. 4A). AtProT2–GFP fusion protein was the positive control for proteins localized at the plasma membrane (Fig. 4B; Grallath et al., 2005). Protoplasts expressing HaPTR4–GFP or GFP–HaPTR4 showed localization at the tonoplast (Fig. 4C). Protoplasts expressing HaPTR5–GFP or GFP–HaPTR5 showed fluorescence in small vesicular structures (Fig. 4D). To investigate the possibility that HaPTR5 localizes to the mitochondrial membrane, co-localization with a mitochondrion-specific dye, Mitotracker (Molecular Probes™, Eugene, OR, USA), was performed; however, no clear co-localization was observed (data not shown). Protoplasts transformed with the HaPTR12–GFP construct showed localization at the plasma membrane (Fig. 4F) while the GFP–HaPTR12 construct showed fluorescence at the plasma membrane as well as some endomembranes (Fig. 4E), possibly due to incomplete targeting.


Nitrogen affects cluster root formation and expression of putative peptide transporters.

Paungfoo-Lonhienne C, Schenk PM, Lonhienne TG, Brackin R, Meier S, Rentsch D, Schmidt S - J. Exp. Bot. (2009)

Localization of fusion proteins of HaPTR with GFP in tobacco (Nicotiana tabacum) protoplasts. Confocal laser scanning microscope pictures (left) and corresponding bright field images (right) of tobacco protoplasts transiently expressing fusion proteins of HaPTRs with GFP or free GFP. Free GFP (A), ProT2–GFP (B), HaPTR4–GFP (C), HaPTR5–GFP (D), GFP–HaPTR12 (E), and HaPTR12–GFP (F). Merged images show GFP fluorescence (green) and chlorophyll fluorescence (red). Protoplast diameter ∼40 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2692012&req=5

fig4: Localization of fusion proteins of HaPTR with GFP in tobacco (Nicotiana tabacum) protoplasts. Confocal laser scanning microscope pictures (left) and corresponding bright field images (right) of tobacco protoplasts transiently expressing fusion proteins of HaPTRs with GFP or free GFP. Free GFP (A), ProT2–GFP (B), HaPTR4–GFP (C), HaPTR5–GFP (D), GFP–HaPTR12 (E), and HaPTR12–GFP (F). Merged images show GFP fluorescence (green) and chlorophyll fluorescence (red). Protoplast diameter ∼40 μm.
Mentions: To demonstrate intracellular localization, GFP fusion proteins were transiently expressed in tobacco protoplasts. Free GFP showed characteristic fluorescence in the cytosol (Fig. 4A). AtProT2–GFP fusion protein was the positive control for proteins localized at the plasma membrane (Fig. 4B; Grallath et al., 2005). Protoplasts expressing HaPTR4–GFP or GFP–HaPTR4 showed localization at the tonoplast (Fig. 4C). Protoplasts expressing HaPTR5–GFP or GFP–HaPTR5 showed fluorescence in small vesicular structures (Fig. 4D). To investigate the possibility that HaPTR5 localizes to the mitochondrial membrane, co-localization with a mitochondrion-specific dye, Mitotracker (Molecular Probes™, Eugene, OR, USA), was performed; however, no clear co-localization was observed (data not shown). Protoplasts transformed with the HaPTR12–GFP construct showed localization at the plasma membrane (Fig. 4F) while the GFP–HaPTR12 construct showed fluorescence at the plasma membrane as well as some endomembranes (Fig. 4E), possibly due to incomplete targeting.

Bottom Line: The results provide further evidence that subgroup II of the NRT1/PTR family contains functional di- and tripeptide transporters.Grown in heathland or hydroponic culture with limiting N supply or starved of nutrients, HaPTR genes had the highest expression in cluster roots and non-cluster roots, and leaf expression increased upon re-supply of ON.It is concluded that formation of cluster roots and expression of PTR are regulated in response to N supply.

View Article: PubMed Central - PubMed

Affiliation: School of Integrative Biology, University of Queensland, QLD, 4072, Australia. chanyarat@uq.edu.au

ABSTRACT
Non-mycorrhizal Hakea actites (Proteaceae) grows in heathland where organic nitrogen (ON) dominates the soil nitrogen (N) pool. Hakea actites uses ON for growth, but the role of cluster roots in ON acquisition is unknown. The aim of the present study was to ascertain how N form and concentration affect cluster root formation and expression of peptide transporters. Hydroponically grown plants produced most biomass with low molecular weight ON>inorganic N>high molecular weight ON, while cluster roots were formed in the order no-N>ON>inorganic N. Intact dipeptide was transported into roots and metabolized, suggesting a role for the peptide transporter (PTR) for uptake and transport of peptides. HaPTR4, a member of subgroup II of the NRT1/PTR transporter family, which contains most characterized di- and tripeptide transporters in plants, facilitated transport of di- and tripeptides when expressed in yeast. No transport activity was demonstrated for HaPTR5 and HaPTR12, most similar to less well characterized transporters in subgroup III. The results provide further evidence that subgroup II of the NRT1/PTR family contains functional di- and tripeptide transporters. Green fluorescent protein fusion proteins of HaPTR4 and HaPTR12 localized to tonoplast, and plasma- and endomembranes, respectively, while HaPTR5 localized to vesicles of unknown identity. Grown in heathland or hydroponic culture with limiting N supply or starved of nutrients, HaPTR genes had the highest expression in cluster roots and non-cluster roots, and leaf expression increased upon re-supply of ON. It is concluded that formation of cluster roots and expression of PTR are regulated in response to N supply.

Show MeSH
Related in: MedlinePlus