Summary Despite the great agricultural and ecological importance of efficient use of urea‐containing nitrogen fertilizers by crops, molecular and physiological identities of urea transport in higher plants have been investigated only in Arabidopsis. We performed short‐time urea‐influx assays which have identified a low‐affinity and high‐affinity (Km of 7.55 μM) transport system for urea‐uptake by rice roots (Oryza sativa). A high‐affinity urea transporter OsDUR3 from rice was functionally characterized here for the first time among crops. OsDUR3 encodes an integral membrane‐protein with 721 amino acid residues and 15 predicted transmembrane domains. Heterologous expression demonstrated that OsDUR3 restored yeast dur3‐mutant growth on urea and facilitated urea import with a Km of c. 10 μM in Xenopus oocytes. Quantitative reverse‐transcription polymerase chain reaction (qPCR) analysis revealed upregulation of OsDUR3 in rice roots under nitrogen‐deficiency and urea‐resupply after nitrogen‐starvation. Importantly, overexpression of OsDUR3 complemented the Arabidopsis atdur3‐1 mutant, improving growth on low urea and increasing root urea‐uptake markedly. Together with its plasma membrane localization detected by green fluorescent protein (GFP)‐tagging and with findings that disruption of OsDUR3 by T‐DNA reduces rice growth on urea and urea uptake, we suggest that OsDUR3 is an active urea transporter that plays a significant role in effective urea acquisition and utilisation in rice.

Ammonium (NH4+) represents a primary nitrogen source for many plants, its effective transport into and between tissues and further assimilation in cells determine greatly plant nitrogen use efficiency. However, biological components involved in NH4+ movement in woody plants are unclear. Here, we report kinetic evidence for cotton NH4+ uptake and molecular identification of certain NH4+ transporters (AMTs) from cotton (Gossypium hirustum). A substrate‐influx assay using 15N‐isotope revealed that cotton possessed a high‐affinity transport system with a Km of 58 μM for NH4+. Sequence analysis showed that GhAMT1.1–1.3 encoded respectively a membrane protein containing 485, 509 or 499 amino acids. Heterologous functionality test demonstrated that GhAMT1.1–1.3 expression mediated NH4+ permeation across the plasma membrane (PM) of yeast and/or Arabidopsis qko‐mutant cells, allowing a growth restoration of both mutants on NH4+. Quantitative PCR measurement showed that GhAMT1.3 was expressed in roots and leaves and markedly up‐regulated by N‐starvation, repressed by NH4+ resupply and regulated diurnally and age‐dependently, suggesting that GhAMT1.3 should be a N‐responsive gene. Importantly, GhAMT1.3 expression in Arabidopsis improved plant growth on NH4+ and enhanced total nitrogen accumulation (∼50% more), conforming with the observation of 2‐fold more NH4+ absorption by GhAMT1.3‐transformed qko plant roots during a 1‐h root influx period. Together with its targeting to the PM and saturated transport kinetics with a Km of 72 μM for NH4+, GhAMT1.3 is suggested to be a high‐affinity NH4+ permease that may play a significant role in cotton NH4+ acquisition and utilization, adding a new member in the plant AMT family.

Many plants grown with low-millimolar concentration of NH 4 + as a sole nitrogen source develop NH 4 +-toxicity symptoms. To date, crucial molecular identities and a 1 | INTRODUCTION The response of plant growth to environmental nitrogen (N) can be characterized, at least in part, by their ability to effectively absorb nitrogenous substances via a spectrum of transport systems, and the ability to sense different N forms and alter the root system

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Dehydration-responsive element-binding proteins (DREBs) belong to members of the AP2/ERF transcription factor superfamily, which has been reported to involve various abiotic-stress responses and tolerance in plants. However, research on the DREB -family is still limited in alfalfa ( Medicago sativa L.), a forage legume cultivated worldwide. The recent genome-sequence release of the alfalfa cultivar “XinJiangDaYe” allowed us to identify 172 DREBs by a multi-step homolog search. The phylogenetic analysis indicated that such MsDREB s could be classified into 5 groups, namely A-1 (56 members), A-2 (39), A-3 (3), A-4 (61) and 13 (A-5 (13), thus adding substantial new members to the DREB -family in alfalfa. Furthermore, a comprehensive survey in silico of conserved motif, gene structure, molecular weight, and isoelectric point (pI) as well as gene expression was conducted. The resulting data showed that, for cold-stress response, 33 differentially expressed MsDREB s were identified with a threshold of Log2-fold > 1, and most of which were transcriptionally upregulated within 48 h during a cold treatment(s). Moreover, the expression profiling of MsDREB s from two ecotypes of alfalfa subspecies i.e. M. sativa ssp. falcata (F56, from a colder region of Central Asia) and M. sativa ssp. sativa (B47, from Near East) revealed that most of the cold-stress responsive MsDREBs exhibited a significantly lower expression in F56, leading to a proposal of the existence of a distinct mechanism(s) for cold tolerance regulated by DREB-related action, which would have been evolved in alfalfa with a genotypic specificity. Additionally, by examining the transcriptome of a freezing-tolerance species ( M. sativa cv. Zhaodong), eight DREBs were found to be implicated in a long-term freezing-stress adaptation with a great potential. Taken together, the current genome-wide identification in alfalfa points to the importance of some MsDREB s in the cold-stress response, providing some promising molecular targets to be functionally characterized for the improvement of cold tolerance in crops including alfalfa.