The Arabidopsis NRG2 Protein Mediates Nitrate Signaling and Interacts with and Regulates Key Nitrate Regulators

Abstract

We show that NITRATE REGULATORY GENE2 (NRG2), which we identified using forward genetics, mediates nitrate signaling in Arabidopsis thaliana. A mutation in NRG2 disrupted the induction of nitrate-responsive genes after nitrate treatment by an ammonium-independent mechanism. The nitrate content in roots was lower in the mutants than in the wild type, which may have resulted from reduced expression of NRT1.1 (also called NPF6.3, encoding a nitrate transporter/receptor) and upregulation of NRT1.8 (also called NPF7.2, encoding a xylem nitrate transporter). Genetic and molecular data suggest that NRG2 functions upstream of NRT1.1 in nitrate signaling. Furthermore, NRG2 directly interacts with the nitrate regulator NLP7 in the nucleus, but nuclear retention of NLP7 in response to nitrate is not dependent on NRG2. Transcriptomic analysis revealed that genes involved in four nitrogen-related clusters including nitrate transport and response to nitrate were differentially expressed in the nrg2 mutants. A nitrogen compound transport cluster containing some members of the NRT/ PTR family was regulated by both NRG2 and NRT1.1, while no nitrogen-related clusters showed regulation by both NRG2 and NLP7. Thus, NRG2 plays a key role in nitrate regulation in part through modulating NRT1.1 expression and may function with NLP7 via their physical interaction.

Whirly1 enhances tolerance to chilling stress in tomato via protection of photosystem II and regulation of starch degradation

“…Here, 5‐wk‐old Nicotiana benthamiana seedlings were used for the transformation as described by Xu et al . (). Fluorescence was measured using a Lumina II instrument (Xenogen, Alameda, CA, USA) and d ‐Luciferin Firefly (Gold Biotechnology, St Louis, MO, USA).…”

Source and sink mechanisms of nitrogen transport and use

“…Nitrate sensing in Arabidopsis involves the transceptor (transporter/receptor) NPF6.3/NRT1.1 that governs many physiological and morphological responses to nitrate such as seed dormancy and lateral root formation (Bouguyon et al, 2015). Nitratesignaling pathways entail calmodulin interacting protein kinases CIPK8 and CIPK23, and the transcription factors SPL9 (Squamosa Promoter Binding Protein-Like 9), TGA1/4 (BZIP proteins) and NLP7 (Nodule Incepetion-Like Protein 7) Krouk et al, 2010a,b;Konishi & Yanagisawa, 2013;Xu et al, 2016). Nitrate triggers the nuclear retention of NLP7, which binds hundreds of genes including many associated with nitrate signaling and assimilation (Marchive et al, 2013;Guan et al, 2017).…”

The long noncoding RNA T5120 regulates nitrate response and assimilation in Arabidopsis

“…Col‐0 was used as the wild‐type (WT) ecotype of Arabidopsis thaliana , and homozygous transgenic seeds carrying a nitrate‐responsive yellow fluorescent protein reporter construct (NRP–YFP), were used as WT for the observation of fluorescence signals (Wang et al ., ). The three nlp7 mutant lines ( nlp7‐1 , nlp7‐2 , and nlp7‐4 containing NRP–YFP) (Castaings et al ., ; Xu et al ., ) and the transgenic lines (p NLP7 ::NLP7−GFP/ nlp7‐1 and p 35S ::NLP7−GFP/ nlp7‐1 ) (Marchive et al ., ) were previously described. The mutant lines chl1‐5 (Ho et al ., ), cipk8‐1 (salk_139697) (Hu et al ., ), nrg2‐2 (salk_079096) (Xu et al ., ), cpsf30‐2 (Li et al ., ) were previously described.…”

“…The three nlp7 mutant lines ( nlp7‐1 , nlp7‐2 , and nlp7‐4 containing NRP–YFP) (Castaings et al ., ; Xu et al ., ) and the transgenic lines (p NLP7 ::NLP7−GFP/ nlp7‐1 and p 35S ::NLP7−GFP/ nlp7‐1 ) (Marchive et al ., ) were previously described. The mutant lines chl1‐5 (Ho et al ., ), cipk8‐1 (salk_139697) (Hu et al ., ), nrg2‐2 (salk_079096) (Xu et al ., ), cpsf30‐2 (Li et al ., ) were previously described. Transgenic lines carrying p 35S :: T5120 were obtained by floral dipping (Clough & Bent, ) of the WT and nlp7‐4 mutant.…”

“…Transcriptional analysis and experimental validation indicate that the expression of > 1000 genes including NRT2.1 , NIA1 , and NIR is altered after nitrate treatment (Wang et al ., , ; Scheible et al ., ; Xu et al ., ; Li et al ., ). Recently, some genes containing NRT1.1 , NLP6/7 , LBD37/38/39 , SPL9 , TGA1/4 , CIPK8/23 , NRG2 , CPSF30‐L , and FIP1 have been identified as important regulators in primary nitrate response (Alvarez et al ., ; Medici & Krouk, ; Xu et al ., ; Wang C. et al ., ; Wang Y. Y. et al ., ; Fredes et al ., ). The long‐term nitrate response is defined as nitrate's effects on aspects of longer term plant growth and development including seed dormancy, root system architecture, the circadian system, and stomatal movement (Roenneberg & Rehman, ; Forde, , ; Guo et al ., ; Walchliu et al ., ; Wilkinson et al ., ; Bisseling & Scheres, ; Guan et al ., ; O'Brien et al ., ).…”