Molecular identification of tobacco NtAMT1.3 that mediated ammonium root-influx with high affinity and improved plant growth on ammonium when overexpressed in Arabidopsis and tobacco

•A functional ammonium transporter NtAMT1.3 was isolated from tobacco.•Expression of NtAMT1.3 complemented the growth on NH4+ of both yeast and Arabidopsis mutant defective in NH4+-uptake.•NtAMT1.3 targets to the plasma membrane and mediated NH4+ root-influx with high affinity at Km of ca. 51 μM.•Transcriptional expression of NtAMT1.3 occurs in both roots and leaves and is upregulated by N-starvation and NH4+.•Overexpression of NtAMT1.3 improved the growth on low NH4+ of tobacco plants.

Although biological functions of ammonium (NH4+) transporters (AMTs) have been intensively studied in many plant species, little is known about molecular bases responsible for NH4+ movement in tobacco. Here, we reported the identification and functional characterization of a putative NH4+ transporter NtAMT1.3 from tobacco (Nicotiana tabacum). Analysis in silico showed that NtAMT1.3 encoded an integral membrane protein containing 464 amino acid residues and exhibiting 10 predicted transmembrane α-helices. Heterologous functionality study demonstrated that NtAMT1.3 expression facilitated NH4+ entry across plasma membrane of NH4+-uptake defective yeast and Arabidopsis qko mutant, allowing a restored growth of both yeast and Arabidopsis mutant on low NH4+. qPCR assay revealed that NtAMT1.3 was expressed in both roots and leaves and significantly up-regulated by nitrogen starvation and resupply of its putative substrate NH4+ and even nitrate, suggesting that NtAMT1.3 should represent a nitrogen-responsive gene. Critically, constitutive overexpression of NtAMT1.3 in tobacco per se improved obviously the growth of transgenic plants on NH4+ and enhanced leaf nitrogen (15% more) accumulation, consistent with observation of 35% more NH4+ uptake by the roots of transgenic lines in 20 min root-influx test. Together with data showing its plasma membrane localization and saturated transport nature with Km of about 50 μM for NH4+, we suggest that NtAMT1.3 acts an active NH4+ transporter that plays a significant role in NH4+ acquisition and utilization in tobacco.