Identification of a 150 bp cis‐acting element of the AtNRT2.1 promoter involved in the regulation of gene expression by the N and C status of the plant

Abstract

The Arabidopsis thaliana AtNRT2.1 gene, which encodes a NO3-transporter involved in high-affinity uptake by the roots, is a molecular target of several mechanisms responsible for the regulation of root NO3 -acquisition by the N status of the plant. All levels of AtNRT2.1 expression (promoter activity, transcript level, protein accumulation, transport activity) are coordinately up-regulated in the presence of NO3 -, and repressed by downstream N metabolites. Transgenic plants expressing the GUS reporter gene under the control of upstream sequences of AtNRT2.1 have been studied to identify elements targeted by these two regulatory mechanisms. A 150 bp sequence located upstream of the TATA box that is required for both stimulation by NO3 -and repression by N metabolites of the promoter has been identified. This sequence is able to confer these two regulations to a minimal promoter. Split-root experiments indicate that the stimulation of the chimaeric promoter by NO3 -occurs only at the local level, whereas its repression by N metabolites is mediated by a systemic signal spread to the whole plant. The activity of the cis-acting 150 bp element is also regulated by sucrose supply to the roots, suggesting a possible interaction between N and C signalling within this short region. Accordingly, multiple motifs potentially involved in regulations by N and/or C status are identified within this sequence by bioinformatic approaches. This is the first report of such a cis-acting element in higher plants.

Nuclear retention of the transcription factor NLP7 orchestrates the early response to nitrate in plants

“…In the case of NIA1, NLP7 binding was also detected downstream of the gene, consistent with the requirement of additional 3 0 end sequences for the proper induction of this gene by nitrate 20 . To directly test the effect of NLP7 on its bound genes, transient transactivation assays were performed in Arabidopsis protoplasts using the reporter construct pNRT2.1:LUC, which contains the luciferase gene under the control of the NRT2.1 promoter 21 and includes the sequence bound by NLP7. Co-transformation of pNRT2.1:LUC with p35S:NLP7 increased the luciferase activity sevenfold compared with the control pNRT2.1:LUC without p35S:NLP7 (Fig.…”

“…For promoter transactivation assays, Arabidopsis protoplasts were prepared from cell cultures by a sucrose gradient after 3 h of cell wall digestion as in Thomine et al 39 polyethylene glycol (PEG)-mediated transfection was performed using 10 mg of plasmid DNA. Plasmids were obtained from M. Lepetit (pNRT2.1:LUC) 21 and from B. Dubreucq (pACT:LEC2) 40 . The p35S:NLP7 construct was obtained by subcloning into pBS 7 .…”

Nitrate foraging by Arabidopsis roots is mediated by the transcription factor TCP20 through the systemic signaling pathway

“…The NIA1 109-bp DNA fragment (16) and the NRT2.1 150-bp DNA fragment (49) were used for screening the transcription factor library as described (59,72). Bait strains were obtained by homologous recombination of pLacZi (Clontech) bait vectors in the yeast YM4271 (Clontech).…”

“…Several nitrate enhancers have been identified in the NRT2.1, NIA1, and NiR genes (49)(50)(51)(52). For NRT2.1, a 150-bp fragment was found that confers nitrate inducibility and N metabolite repression on a minimal core promoter (49).…”

Identification of Arabidopsis Mutants Impaired in the Systemic Regulation of Root Nitrate Uptake by the Nitrogen Status of the Plant

“…Similar results were obtained with the APSR gene, encoding 5#-adenosine phosphosulfate reductase (At1g62180; data not shown). This suggested that 2 as sole N source according to Girin et al (2007). Values are means of six replicates 6 SD.…”