Isolation and characterization of a high-affinity ammonium transporter ApAMT1;1 in alligatorweed

Abstract

In aquatic fields, ammonium (NH4+) is the most preferred nitrogen (N) source used by plants. The uptake of NH4+ is facilitated by the family of ammonium transporters (AMTs). However, the molecular functions of AMTs in aquatic plants are largely unknown. In this work, a new NH4+ transporter encoding gene, ApAMT1;1, was isolated from the typical aquatic plant alligatorweed, using degenerated primers and rapid amplification of cDNA end (RACE) techniques. Quantitative real time PCR showed that ApAMT1;1 was predominantly expressed in roots, and significantly induced by NH4+ starvation in all tested tissues, including leaves, stems and roots. Functional determination and 15N-labeled ammonium uptake assays in yeast cells indicated that ApAMT1;1 was a typical high-affinity transporter, with a 38.6 μM Km value, and the phosphorylation site T469 was required to retain its NH4+ uptake capacity. Further analyses with Met sulfoximine (MSX), a NH4+ assimilation inhibitor, demonstrated that ApAMT1;1-mediated NH4+ uptake might be feedback regulated by the internal NH4+ accumulation. Our results reveal a functional role of ApAMT1;1 in the uptake and transport of NH4+ in aquatic plants.