A 150 kDa plasma membrane complex of AtNRT2.5 and AtNAR2.1 is the major contributor to constitutive high‐affinity nitrate influx in Arabidopsis thaliana

Abstract

In plants that have been deprived of nitrate for a significant length of time, a constitutive high-affinity nitrate transport system (cHATS) is responsible for initial nitrate uptake. This absorbed nitrate leads to the induction of the major nitrate transporters and enzymes involved in nitrate assimilation. By use of (13) NO3 (-) influx measurements and Blue Native polyacrylamide gel electrophoresis we examined the role of AtNRT2.5 in cHATS in wild type (WT) and various T-DNA mutants of Arabidopsis thaliana. We demonstrate that AtNRT2.5 is predominantly expressed in roots of nitrate-deprived WT plants as a 150 kDa molecular complex with AtNAR2.1. This complex represents the major contributor to cHATS influx, which is reduced by 63% compared with WT in roots of Atnrt2.5 mutants. The remaining cHATS nitrate influx in these mutants is due to a residual contribution by the inducible high-affinity transporter encoded by AtNRT2.1/AtNAR2.1. Estimates of the kinetic properties of the NRT2.5 transporter reveal that its low Km for nitrate makes this transporter ideally suited to detect and respond to trace quantities of nitrate in the root environment.

Time-Resolved Investigation of Molecular Components Involved in the Induction of NO3– High Affinity Transport System in Maize Roots

“…Interactions between members of NRT2 family with NRT3 (NAR2) proteins were reported in Arabidopsis (Orsel et al, 2006; Yong et al, 2010; Kotur et al, 2012; Kotur and Glass, 2015), rice (Yan et al, 2011; Liu et al, 2014), barley (Ishikawa et al, 2009), and Chrysanthemum morifolium (Gu et al, 2016). These proteins appear to be associated in an oligomer of ∼150 kDa as observed in Arabidopsis roots in response to ${\mathrm{NO}}_{\mathrm{3}}^{\mathrm{–}}$ induction, thus suggesting that iHATS relies on a tetrameric aggregation of two subunits, namely AtNRT2.1 and AtNAR2.1 (Yong et al, 2010).…”

“…These proteins appear to be associated in an oligomer of ∼150 kDa as observed in Arabidopsis roots in response to ${\mathrm{NO}}_{\mathrm{3}}^{\mathrm{–}}$ induction, thus suggesting that iHATS relies on a tetrameric aggregation of two subunits, namely AtNRT2.1 and AtNAR2.1 (Yong et al, 2010). Recently, a 150 kDa complex of AtNRT2.5 and AtNAR2.1 was shown to be involved in constitutive HATS (cHATS; Kotur and Glass, 2015). …”

Plant nitrogen assimilation and its regulation: a complex puzzle with missing pieces

“…Under low-nitrate conditions, the main transport activity is achieved by AtNRT2.1, whereas AtNRT2.2 makes only a small contribution [8]. AtNRT2.4 and AtNRT2.5 are highly induced by N starvation and are expressed in the root epidermis [9][10][11]. Whereas AtNRT2.1 loss of function reduces plant growth under low external nitrate, growth is not particularly restricted by loss of either AtNRT2.2, AtNRT2.4 or AtNRT2.5.…”

“…Hsu PK, Tsay YF: Two phloem nitrate transporters, NRT1 11. and NRT1.12, are important for redistributing xylem-borne nitrate to enhance plant growth.…”

Environmental nitrate signals through abscisic acid in the root tip

“…6,20 Under conditions of low external nitrate, uptake is mediated primarily by the high-affinity transporter, AtNRT2.1, and to a lesser degree by AtNRT2.2 and AtNRT2.4 [21][22][23] ; under nitrate starvation conditions uptake is mediated mainly by AtNRT2.5, with some contribution by AtNRT2.1. 24,25 At high concentrations of environmental nitrate, the dual-affinity transporter AtNPF6.3 (AtNRT1.1) and the low-affinity transporter AtNPF4.6 (AtNRT1.2; AtAIT1) come into play. 26,27 Two of these transporters, AtNPF6.3 and AtNRT2.1, have also been proposed to function as receptors, thus earning the name of "transceptor".…”