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No Uptake Mediated by Zosmanrt2 Is A Na - Dependent Mechanism

This study characterized the function of ZosmaNRT2, a nitrate transporter gene from the seagrass Zostera marina. The researchers expressed ZosmaNRT2 in the yeast Hansenula polymorpha, which lacks its own high-affinity nitrate transporter. They found that nitrate uptake mediated by ZosmaNRT2 requires sodium ions, unlike other plant NRT2 transporters which rely on protons. This provides the first evidence of a sodium-dependent nitrate uptake system in the angiosperm NRT2 family and establishes a functional model for this novel transport mechanism.

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jordi D. G.
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30 views1 page

No Uptake Mediated by Zosmanrt2 Is A Na - Dependent Mechanism

This study characterized the function of ZosmaNRT2, a nitrate transporter gene from the seagrass Zostera marina. The researchers expressed ZosmaNRT2 in the yeast Hansenula polymorpha, which lacks its own high-affinity nitrate transporter. They found that nitrate uptake mediated by ZosmaNRT2 requires sodium ions, unlike other plant NRT2 transporters which rely on protons. This provides the first evidence of a sodium-dependent nitrate uptake system in the angiosperm NRT2 family and establishes a functional model for this novel transport mechanism.

Uploaded by

jordi D. G.
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Functional characterization of ZosmaNRT2 by heterologous expression in Hansenula polymorpha

Jordi Díaz-García1, José M. Siverio2, Javier Quintero3, Lourdes Rubio1, José A. Fernández1
1Dpt. Botánica y Fisiología Vegetal, UMA. 2Dpt. Bioquímica, Microbiología, Biología Celular y Genética, ULL. 3Instituto de Bioquímica Vegetal y Fotosíntesis, CSIC, Sevilla.

([email protected])

Zostera marina L. is a seagrass, the unique vascular plant group that colonized the marine environment. This involved significant evolutionary challenges to thrive in liquid medium containing high salinity, alkaline pH
and very low NO3- concentrations. In contrast to vascular land plants, only one sequence (ZosmaNRT2) belonging to the high-affinity NO3- transporters NRT2 was found at the Z. marina genome, which was proposed to
encode the putative Na+-dependent high-affinity NO3- transporter of this seagrass (1). In this work, heterologous expression in Hansenula polymorpha Δynt1, a high-affinity nitrate transporter deficient strain, is used to
unveil the ZosmaNTRT2 transport mechanism.
NO3- uptake mediated by ZosmaNRT2 is a Na+ -dependent mechanism
ZosmaNRT2 structural analysis predicts a high homology with AtNRT2.5, showing the same and single NO3- binding site, but no prediction for the driving ion binding site was found (Fig. 1). However, in H. polymorpha
complement strain, expressing ZosmaNRT2, NO3- uptake requires Na+ instead H+, which are considered the driving ion for NO3- uptake mediated by NRT2 transporters and also by ∆YNT1 (Fig. 2, Table 1). Thus, results
presented here are the first functional evidence of a Na+-dependent NO3- uptake system belonging to the angiosperm NRT2 family.
A B C D E
Table 1. NO3- depletion rates calculated as the initial slope of the depletion curves in
Fig. 2. Values were normalized to the cells wet weight. Data are mean ± SD (n = 3).
Different letters denote significant differences (2-way ANOVA, Tukey's test, p <0.05).

Fig. 1. Predicted NRT2 3D Structure. ZosmaNRT2 (A), AtNRT2.5 (B) are shown in blue and red, respectively.
Superimposed proteins (C). Top view of predicted NO3- -binding site (D) and amino acid composition: PHE65, ARG104,
PHE157, ASN185, TYR277, PHE281, ARG321 (E). Dotted lines denote distance between atoms. Models obtained from
homology/analogy recognition engines I-TASSER.

0.5M Na+ pH 8.2


< 10 μM NO3-

Na+

Fig. 2. Nitrate depletion assays was performed using 20 mg of cells (wet weight) in 1 mL of synthetic N-free yeast base
(0.17% w/v) supplemented with 25 mM NaCl or 50 mM Sorbitol and buffered at pH 6.3 or 8.3 using 20 mM MES/BTP. ZosmaNRT2
After the initial addition of 50 µM KNO3, external NO3- concentration was colorimetrically monitored at different times First functional model for a Na+-dependent NO3- uptake mechanism
(2). Data are mean±SD (n=3).
in angiosperms
(1) Rubio et al., 2019. International Journal Molecular Science 20,3650.
(2) Robledo et al., 2014. Marine Chemistry 162:30-36 BFU2017-85117-R; BIO2016-81957-REDT; FPU18/03300

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