NIN-like Protein Research Articles

The AP2/ERF Transcription Factor ERF56 Negatively Regulating Nitrate-Dependent Plant Growth in Arabidopsis.

ERF56, a member of the APETALA2/ETHYLENE-RESPONSIVE FACTOR (AP2/ERF) transcription factor (TF) family, was reported to be an early nitrate-responsive TF in Arabidopsis. But the function of ERF56 in nitrate signaling remains not entirely clear. This study aimed to investigate the role of ERF56 in nitrate-dependent plant growth and nitrate signaling. We confirmed with reverse transcription quantitative PCR (RT-qPCR) that the transcription of ERF56 is quickly induced by nitrate. ERF56 overexpressors displayed decreased nitrate-dependent plant growth, while erf56 mutants exhibited increased plant growth. Confocal imaging demonstrated that ERF56 is localized into nuclei. Assays with the glucuronidase (GUS) reporter showed that ERF56 is mainly expressed at the region of maturation of roots and in anthers. The dual-luciferase assay manifested that the transcription of ERF56 is not directly regulated by NIN-LIKE PROTEIN 7 (NLP7). The transcriptome analysis identified 1038 candidate genes regulated by ERF56 directly. A gene ontology (GO) over-representation analysis showed that ERF56 is involved in the processes of water transport, inorganic molecule transmembrane transport, secondary metabolite biosynthesis, and cell wall organization. We revealed that ERF56 represses nitrate-dependent growth through regulating the processes of inorganic molecule transmembrane transport, the secondary metabolite biosynthesis, and cell wall organization.

Carbon and nitrogen signaling regulate FLOWERING LOCUS C and impact flowering time in Arabidopsis.

The timing of flowering in plants is modulated by both carbon (C) and nitrogen (N) signaling pathways. In a previous study, we established a pivotal role of the sucrose-signaling trehalose 6-phosphate pathway in regulating flowering under N-limited short-day conditions. In this work, we show that both wild-type Arabidopsis (Arabidopsis thaliana) plants grown under N-limited conditions and knock-down plants of TREHALOSE PHOSPHATE SYNTHASE 1 induce FLOWERING LOCUS C (FLC) expression, a well-known floral repressor associated with vernalization. When exposed to an extended period of cold, a flc mutant fails to respond to N availability and flowers at the same time under N-limited and full-nutrition conditions. Our data suggest that SUCROSE NON-FERMENTING 1 RELATED KINASE 1-dependent trehalose 6-phosphate-mediated C signaling and a mechanism downstream of N signaling (likely involving NIN-LIKE PROTEIN 7) impact the expression of FLC. Collectively, our data underscore the existence of a multi-factor regulatory system in which the C and N signaling pathways jointly govern the regulation of flowering in plants.

High-nitrogen-induced γ-aminobutyric acid triggers host immunity and pathogen oxidative stress tolerance in tomato and Ralstonia solanacearum interaction.

Soil nitrogen (N) significantly influences the interaction between plants and pathogens, yet its impact on host defenses and pathogen strategies via alterations in plant metabolism remains unclear. Through metabolic and genetic studies, this research demonstrates that high-N-input exacerbates tomato bacterial wilt by altering γ-aminobutyric acid (GABA) metabolism of host plants. Under high-N conditions, the nitrate sensor NIN-like protein 7 (SlNLP7) promotes the glutamate decarboxylase 2/4 (SlGAD2/4) transcription and GABA synthesis by directly binding to the promoters of SlGAD2/4. The tomato plants with enhanced GABA levels showed stronger immune responses but remained susceptible to Ralstonia solanacearum. This led to the discovery that GABA produced by the host actually heightens the pathogen's virulence. We identified the R. solanacearum LysR-type transcriptional regulator OxyR protein, which senses host-derived GABA and, upon interaction, triggers a response involving protein dimerization that enhances the pathogen's oxidative stress tolerance by activating the expression of catalase (katE/katGa). These findings reveal GABA's dual role in activating host immunity and enhancing pathogen tolerance to oxidative stress, highlighting the complex relationship between tomato plants and R. solanacearum, influenced by soil N status.

Mapping the molecular landscape of Lotus japonicus nodule organogenesis through spatiotemporal transcriptomics

Legumes acquire nitrogen-fixing ability by forming root nodules. Transferring this capability to more crops could reduce our reliance on nitrogen fertilizers, thereby decreasing environmental pollution and agricultural production costs. Nodule organogenesis is complex, and a comprehensive transcriptomic atlas is crucial for understanding the underlying molecular events. Here, we utilized spatial transcriptomics to investigate the development of nodules in the model legume, Lotus japonicus. Our investigation has identified the developmental trajectories of two critical regions within the nodule: the infection zone and peripheral tissues. We reveal the underlying biological processes and provide gene sets to achieve symbiosis and material exchange, two essential aspects of nodulation. Among the candidate regulatory genes, we illustrate that LjNLP3, a transcription factor belonging to the NIN-LIKE PROTEIN family, orchestrates the transition of nodules from the differentiation to maturation. In summary, our research advances our understanding of nodule organogenesis and provides valuable data for developing symbiotic nitrogen-fixing crops.

The beneficial rhizobacterium Bacillus velezensis SQR9 regulates plant nitrogen uptake via an endogenous signaling pathway.

Nitrogen fertilizer is widely used in agriculture to boost crop yields. Plant growth-promoting rhizobacteria (PGPRs) can increase plant nitrogen use efficiency through nitrogen fixation and organic nitrogen mineralization. However, it is not known whether they can activate plant nitrogen uptake. In this study, we investigated the effects of volatile compounds (VCs) emitted by the PGPR strain Bacillus velezensis SQR9 on plant nitrogen uptake. Strain SQR9 VCs promoted nitrogen accumulation in both rice and Arabidopsis. In addition, isotope labeling experiments showed that strain SQR9 VCs promoted the absorption of nitrate and ammonium. Several key nitrogen-uptake genes were up-regulated by strain SQR9 VCs, such as AtNRT2.1 in Arabidopsis and OsNAR2.1, OsNRT2.3a, and OsAMT1 family members in rice, and the deletion of these genes compromised the promoting effect of strain SQR9 VCs on plant nitrogen absorption. Furthermore, calcium and the transcription factor NIN-LIKE PROTEIN 7 play an important role in nitrate uptake promoted by strain SQR9 VCs. Taken together, our results indicate that PGPRs can promote nitrogen uptake through regulating plant endogenous signaling and nitrogen transport pathways.

Genome-wide identification and expression analysis of NIN-like protein (NLP) genes: Exploring their potential roles in nitrate response in tea plant (Camellia sinensis)

NIN-like proteins (NLPs) are evolutionarily conserved transcription factors that are unique to plants and play a pivotal role in responses to nitrate uptake and assimilation. However, a comprehensive analysis of NLP members in tea plants is lacking. The present study performed a genome-wide analysis and identified 33 NLP gene family members of Camellia sinensis that were distributed unequally across 5 chromosomes. Subcellular localisation predictions revealed that all CsNLP proteins were localised in the nucleus. Conservative domain analysis revealed that all of these proteins contained conserved RWP-RK and PB1 domains. Phylogenetic analysis grouped the CsNLP gene family into four clusters. The promoter regions of CsNLPs harboured cis-acting elements associated with plant hormones and abiotic stress responses. Expression profile analysis demonstrated that CsNLP8 was significantly upregulated in roots under nitrate stress conditions. Subcellular localisation analysis found CsNLP8 localised to the nucleus. Dual-luciferase reporter assay demonstrated that CsNLP8 positively regulated the expression of a nitrate transporter gene (CsNRT2.2). These findings provide a comprehensive characterisation of NLP genes in Camellia sinensis and offer insights into the biological function of CsNLP8 in regulating the response to nitrate-induced stress.

The NIN-LIKE PROTINE1 (CsNLP1) transcription factor is involved in modulating the nitrate response in cucumber seedlings

Cucumber is one of the important vegetables cultivated in facilities in China. Due to the deterioration of soil quality in facilities and the low nitrogen use efficiency of existing varieties, it is common to pursue high yield through excessive application of nitrogen fertilizer. Improving crop nitrogen use efficiency is one of the important means to reduce nitrogen fertilizer input. Previous studies have shown that NLP (NIN-like protein) transcription factor family play an important role as transcription factors in regulating plant physiological responses to external nitrogen changes. However, there are few reports on NLPs in cucumber. Here we report that CsNLP1 is a potential candidate to improve cucumber nitrogen use ability. CsNLP1 silencing lines by virus-induced gene silencing (VIGS) showed a significant decrease in the plant biomass, chlorophyll content, nitrogen uptake, total nitrogen content, activities of NR, NIR, GDH, GOGAT and GS, and expression levels of genes involved in nitrogen assimilation and signalling under different nitrogen concentrations. In addition, transfer of CsNLP1 into Arabidopsis nlp7-1 mutant found that the phenotype, nitrate (NO3-) content, NR activity, NIR activity, GDH activity, GOGAT activity and GS activity in transgenic lines could be restored to the wild type level after NO3- treatment. Further study found that CsNLP1 could regulate the expression of NO3- responsive genes such as AtNIR1, AtNIA2, AtNRT1.1 and AtGS2 in plants. Overall, our data indicated that CsNLP1 can affect plant growth, and regulate nitrate assimilation by regulating nitrogen-related genes.

Calcium regulates primary nitrate responseassociated gene transcription in a time- and dose-dependent manner.

Nitrate (NO3-) is the primary source of nitrogen preferred by most arable crops, including wheat. The pioneering experiment on primary nitrate response (PNR) was carried out three decades ago. Since then, much research has been carried out to understand the NO3- signaling. Nitrate is sensed by the dual affinity NO3- transceptor NPF6.3, which further relays the information to a master regulator NIN-like protein 7 (NLP7) through calcium-dependent protein kinases (CPK10, CPK30, CPK32), highlighting the importance of calcium ion (Ca2+) as one of the important secondary messengers in relaying the NO3- signaling in Arabidopsis. In a previous study, we found that Ca2+ regulates nitrogen starvation response in wheat. In this study, 10days old NO3--starved wheat seedlings were exposed to various treatments. Our study on time course changes in expression of PNR sentinel genes; NPF6.1, NPF6.2, NRT2.1, NRT2.3, NR, and NIR in wheat manifest the highest level of expression at 30min after NO3- exposure. The use of Ca2+ chelator EGTA confirmed the involvement of Ca2+ in the regulation of transcription of NPFs and NRTs as well the NO3- uptake. We also observed the NO3- dose-dependent and tissue-specificregulation of nitrate reductase activity involving Ca2+ as a mediator. The participation of Ca2+ in the PNR and NO3- signaling in wheat is confirmed by pharmacological analysis, physiological evidences, and protoplast-based Ca2+ localization.

PIF4 enhances the expression of SAUR genes to promote growth in response to nitrate

Nitrate supply is fundamental to support shoot growth and crop performance, but the associated increase in stem height exacerbates the risks of lodging and yield losses. Despite their significance for agriculture, the mechanisms involved in the promotion of stem growth by nitrate remain poorly understood. Here, we show that the elongation of the hypocotyl of Arabidopsis thaliana, used as a model, responds rapidly and persistently to upshifts in nitrate concentration, rather than to the nitrate level itself. The response occurred even in shoots dissected from their roots and required NITRATE TRANSPORTER 1.1 (NRT1.1) in the phosphorylated state (but not NRT1.1 nitrate transport capacity) and NIN-LIKE PROTEIN 7 (NLP7). Nitrate increased PHYTOCHROME INTERACTING FACTOR 4 (PIF4) nuclear abundance by posttranscriptional mechanisms that depended on NRT1.1 and phytochrome B. In response to nitrate, PIF4 enhanced the expression of numerous SMALL AUXIN-UP RNA (SAUR) genes in the hypocotyl. The growth response to nitrate required PIF4, positive and negative regulators of its activity, including AUXIN RESPONSE FACTORs, and SAURs. PIF4 integrates cues from the soil (nitrate) and aerial (shade) environments adjusting plant stature to facilitate access to light.

Balanced nitrogen–iron sufficiency boosts grain yield and nitrogen use efficiency by promoting tillering

Crop yield plays a critical role in global food security. For optimal plant growth and maximal crop yields, nutrients must be balanced. However, the potential significance of balanced nitrogen–iron (N–Fe) for improving crop yield and nitrogen use efficiency (NUE) has not previously been addressed. Here, we show that balanced N–Fe sufficiency significantly increases tiller number and boosts yield and NUE in rice and wheat. NIN-like protein 4 (OsNLP4) plays a pivotal role in maintaining the N–Fe balance by coordinately regulating the expression of multiple genes involved in N and Fe metabolism and signaling. OsNLP4 also suppresses OsD3 expression and strigolactone (SL) signaling, thereby promoting tillering. Balanced N–Fe sufficiency promotes the nuclear localization of OsNLP4 by reducing H2O2 levels, reinforcing the functions of OsNLP4. Interestingly, we found that OsNLP4 upregulates the expression of a set of H2O2-scavenging genes to promote its own accumulation in the nucleus. Furthermore, we demonstrated that foliar spraying of balanced N–Fe fertilizer at the tillering stage can effectively increase tiller number, yield, and NUE of both rice and wheat in the field. Collectively, these findings reveal the previously unrecognized effects of N–Fe balance on grain yield and NUE as well as the molecular mechanism by which the OsNLP4–OsD3 module integrates N–Fe nutrient signals to downregulate SL signaling and thereby promote rice tillering. Our study sheds light on how N–Fe nutrient signals modulate rice tillering and provide potential innovative approaches that improve crop yield with reduced N fertilizer input for benefitting sustainable agriculture worldwide.

Transcription factor module NLP-NIGT1 fine-tunes NITRATE TRANSPORTER2.1 expression.

Arabidopsis (Arabidopsis thaliana) high-affinity NITRATE TRANSPORTER2.1 (NRT2.1) plays a dominant role in the uptake of nitrate, the most important nitrogen (N) source for most terrestrial plants. The nitrate-inducible expression of NRT2.1 is regulated by NIN-LIKE PROTEIN (NLP) family transcriptional activators and NITRATE-INDUCIBLE GARP-TYPE TRANSCRIPTIONAL REPRESSOR1 (NIGT1) family transcriptional repressors. Phosphorus (P) availability also affects the expression of NRT2.1 because the PHOSPHATE STARVATION RESPONSE1 transcriptional activator activates NIGT1 genes in P-deficient environments. Here, we show a biology-based mathematical understanding of the complex regulation of NRT2.1 expression by multiple transcription factors using 2 different approaches: a microplate-based assay for the real-time measurement of temporal changes in NRT2.1 promoter activity under different nutritional conditions, and an ordinary differential equation (ODE)-based mathematical modeling of the NLP- and NIGT1-regulated expression patterns of NRT2.1. Both approaches consistently reveal that NIGT1 stabilizes the amplitude of NRT2.1 expression under a wide range of nitrate concentrations. Furthermore, the ODE model suggests that parameters such as the synthesis rate of NIGT1 mRNA and NIGT1 proteins and the affinity of NIGT1 proteins for the NRT2.1 promoter substantially influence the temporal expression patterns of NRT2.1 in response to nitrate. These results suggest that the NLP-NIGT1 feedforward loop allows a precise control of nitrate uptake. Hence, this study paves the way for understanding the complex regulation of nutrient acquisition in plants, thus facilitating engineered nutrient uptake and plant response patterns using synthetic biology approaches.

Identification and expression characteristics of NLP (NIN-like protein) gene family in pepper (Capsicum annuum L.).

Pepper (Capsicum annum L.) is the main crop in the vegetable industry. The growth and development of peppers are regulated by nitrate, but there is limited research on the molecular mechanisms of nitrate absorption and assimilation in peppers. A plant specific transcription factor NLP plays an important role in nitrate signal transduction. In this study, a total of 7 NLP members were identified based on pepper genome data. Two nitrogen transport elements (GCN4) were found in the CaNLP5 promoter. In the phylogenetic tree, CaNLP members are divided into three branches, with pepper NLP and tomato NLP having the closest genetic relationship. The expression levels of CaNLP1, CaNLP3, and CaNLP4 are relatively high in the roots, stems, and leaves. The expression level of CaNLP7 gene is relatively high during the 5-7days of pepper fruit color transformation. After various non-Biotic stress and hormone treatments, the expression of CaNLP1 was at a high level. The expression of CaNLP3 and CaNLP4 was down regulated in leaves, but up regulated in roots. Under conditions of nitrogen deficiency and sufficient nitrate, the expression patterns of NLP genes in pepper leaves and roots were determined. These results provide important insights into the multiple functions of CaNLPs in regulating nitrate absorption and transport.

Genome-Wide Identification and Expression Analysis of AS2 Genes in Brassica rapa Reveal Their Potential Roles in Abiotic Stress

The ASYMMETRIC LEAVES2/LATERAL ORGAN BOUNDARIES (AS2/LOB) gene family plays a pivotal role in plant growth, induction of phytohormones, and the abiotic stress response. However, the AS2 gene family in Brassica rapa has yet to be investigated. In this study, we identified 62 AS2 genes in the B. rapa genome, which were classified into six subfamilies and distributed across 10 chromosomes. Sequence analysis of BrAS2 promotors showed that there are several typical cis-elements involved in abiotic stress tolerance and stress-related hormone response. Tissue-specific expression analysis showed that BrAS2-47 exhibited ubiquitous expression in all tissues, indicating it may be involved in many biological processes. Gene expression analysis showed that the expressions of BrAS2-47 and BrAS2-10 were significantly downregulated under cold stress, heat stress, drought stress, and salt stress, while BrAS2-58 expression was significantly upregulated under heat stress. RT-qPCR also confirmed that the expression of BrAS2-47 and BrAS2-10 was significantly downregulated under cold stress, drought stress, and salt stress, and in addition BrAS2-56 and BrAS2-4 also changed significantly under the three stresses. In addition, protein–protein interaction (PPI) network analysis revealed that the Arabidopsis thaliana genes AT5G67420 (homologous gene of BrAS2-47 and BrAS2-10) and AT3G49940 (homologous gene of BrAS2-58) can interact with NIN-like protein 7 (NLP7), which has been previously reported to play a role in resistance to adverse environments. In summary, our findings suggest that among the BrAS2 gene family, BrAS2-47 and BrAS2-10 have the most potential for the regulation of abiotic stress tolerance. These results will facilitate future functional investigations of BrAS2 genes in B. rapa.

GmNLP7a inhibits soybean nodulation by interacting with GmNIN1a

Nitrogen (N) is an essential macronutrient for plant growth and productivity. Leguminous plants establish symbiotic relationships with nitrogen-fixing rhizobial bacteria to use atmospheric dinitrogen gas to meet high N demand under low-N conditions. Nodule formation and N fixation are energy-consuming processes and are inhibited by nitrate present in the environment. Previous studies in model leguminous plants characterized NIN-LIKE PROTEIN (NLP) proteins that mediate nitrate control of root nodule symbiosis, but the mechanism by which nitrate regulates soybean root nodules via NLP remains unclear. In the soybean genome we found four homologs of AtNLP7, named GmNLP7a–GmNLP7d. We showed that the expression of GmNLP7s is responsive to nitrate but not to rhizobial infection and localized GmNLP7a to the nucleus. Downregulation of GmNLP7s increased nodule number, and overexpression of GmNLP7a (GmNLP7aOE) reduced nodule number regardless of nitrate availability, suggesting a negative role for GmNLP7s in nodulation. Nitrogenase activity in the GmNLP7aOE line was comparable to that of the wild type, indicating that GmNLP7a does not affect mature nodule activity. Overexpression of GmNLP7a downregulated the expression of GmNIN1a and GmENOD40-1. GmNLP7a interacted with GmNIN1a via the PB1 domain. Our results reveal a new regulator of GmNLP7 in nodulation and a molecular mechanism by which nitrate affects nodule number in soybean.

Physiological traits and expression profile of genes associated with nitrogen and phosphorous use efficiency in wheat

Nitrogen (N) and phosphorous (P) play a very important role in the growth and development of wheat as well as major constituents of biological membranes. To meet the plant's nutritional demand these nutrients are applied in the form of fertilizers. But the plant can utilize only half of the applied fertilizer whereas the rest is lost through surface runoff, leaching and volatilization. Thus, to overcome the N/P loss we need to elucidate the molecular mechanism behind the N/P uptake. In our study, we used DBW16 (low NUE), and WH147 (high NUE) wheat genotypes under different doses of N, whereas HD2967 (low PUE) and WH1100 (high PUE) genotypes were studied under different doses of P. To check the effect of different doses of N/P, the physiological parameters like total chlorophyll content, net photosynthetic rate, N/P content, and N/PUE of these genotypes were calculated. In addition, gene expression of various genes involved in N uptake, utilization, and acquisition such as Nitrite reductase (NiR), Nitrate transporter 1/Peptide transporter family (NPF2.4/2.5), Nitrate transporter (NRT1) and NIN Like Protein (NLP) and induced phosphate starvation (IPS), Phosphate Transporter (PHT1.7) and Phosphate 2 (PHO2) acquisition was studied by quantitative real-time PCR. Statistical analysis revealed a lower percent reduction in TCC, NPR, and N/P content in N/P efficient wheat genotypes (WH147 & WH1100). A significant increase in relative fold expression of genes under low N/P concentration was observed in N/P efficient genotypes as compared to N/P deficient genotypes. Significant differences in physiological data and gene expression among N/ P efficient and deficient wheat genotypes could be useful for future improvement of N/P use efficiency.

GmTCP and GmNLP Underlying Nodulation Character in Soybean Depending on Nitrogen.

Soybean is a cereal crop with high protein and oil content which serves as the main source of plant-based protein and oil for human consumption. The symbiotic relationship between legumes and rhizobia contributes significantly to soybean yield and quality, but the underlying molecular mechanisms remain poorly understood, hindering efforts to improve soybean productivity. In this study, we conducted a transcriptome analysis and identified 22 differentially expressed genes (DEGs) from nodule-related quantitative trait loci (QTL) located in chromosomes 12 and 19. Subsequently, we performed functional characterisation and haplotype analysis to identify key candidate genes among the 22 DEGs that are responsive to nitrate. Our findings identified GmTCP (TEOSINTE-BRANCHED1/CYCLOIDEA/PCF) and GmNLP (NIN-LIKE PROTEIN) as the key candidate genes that regulate the soybean nodule phenotype in response to nitrogen concentration. We conducted homologous gene mutant analysis in Arabidopsis thaliana, which revealed that the homologous genes of GmTCP and GmNLP play a vital role in regulating root development in response to nitrogen concentration. We further performed overexpression and gene knockout of GmTCP and GmNLP through hairy root transformation in soybeans and analysed the effects of GmTCP and GmNLP on nodulation under different nitrogen concentrations using transgenic lines. Overexpressing GmTCP and GmNLP resulted in significant differences in soybean hairy root nodulation phenotypes, such as nodule number (NN) and nodule dry weight (NDW), under varying nitrate conditions. Our results demonstrate that GmTCP and GmNLP are involved in regulating soybean nodulation in response to nitrogen concentration, providing new insights into the mechanism of soybean symbiosis establishment underlying different nitrogen concentrations.

The mechanisms of optimal nitrogen conditions to accelerate flowering of Chrysanthemum vestitum under short day based on transcriptome analysis

Nitrogen (N) plays an important role in the development of plants, with N application having been shown to accelerate flowering of cultivated plants. However, the mechanism of optimal N conditions to accelerate flowering of short-day plants is still unclear. In this study, it was found that Chrysanthemum vestitum is a typical short-day plant like most chrysanthemum varieties, and its flowering must go through a short-day induction stage. Further observations on the growth of C. vestitum showed that the N range of external application for growth was limited to between 0.25 and 2.50 mM. The results showed that, under optimal N (ON, 1.25 mM) conditions, the plants increased rapidly and flowering time was advanced; under high N (HN, 2.50 mM) or limited N (LN, 0.25 mM) conditions, the growth of plants were inhibited and flowering time was delayed. On the basis of transcriptome data, analysis of differentially expressed genes (DEGs) revealed that the floral-related genes B-box19 (BBX19), Cryptochromes (CRYs), CONSTANS-like (COLs), nitrate transporter protein (NRT), and NIN-like protein (NLP) could respond to N availability. Most of the genes in the photoperiod pathway were upregulated by ON conditions, and their expression was inhibited under HN and LN conditions. Our findings indicated that N could affect flowering by regulating the transcription levels of genes that are involved mainly in the photoperiod pathway. These candidate genes provide important clues for the subsequent analysis of the mechanism of N-induced flowering of short-day plants, and provide a possibility to improve the flowering of chrysanthemum by molecular breeding.

The NIN-LIKE PROTEIN 7 transcription factor modulates auxin pathways to regulate root cap development in Arabidopsis.

The root cap is a small tissue located at the tip of the root with critical functions for root growth. Present in nearly all vascular plants, the root cap protects the root meristem, influences soil penetration, and perceives and transmits environmental signals that are critical for root branching patterns. To perform these functions, the root cap must remain relatively stable in size and must integrate endogenous developmental pathways with environmental signals, yet the mechanism is not clear. We previously showed that low pH conditions altered root cap development, and these changes are mediated by the NIN LIKE PROTEIN 7 (NLP7) transcription factor, a master regulator of nitrate signaling. Here we show that in Arabidopsis NLP7 integrates nitrate signaling with auxin pathways to regulate root cap development. We found that low nitrate conditions promote aberrant release of root cap cells. Nitrate deficiency impacts auxin pathways in the last layer of the root cap, and this is mediated in part by NLP7. Mutations in NLP7 abolish the auxin minimum in the last layer of the root cap and alter root cap expression of the auxin carriers PIN-LIKES 3 (PILS3) and PIN-FORMED 7 (PIN7) as well as transcription factors that regulate PIN expression. Together, our data reveal NLP7 as a link between endogenous auxin pathways and nitrate signaling in the root cap.

The small peptide CEP1 and the NIN-like protein NLP1 regulate NRT2.1 to mediate root nodule formation across nitrate concentrations.

Legumes acquire fixed nitrogen (N) from the soil and through endosymbiotic association with diazotrophic bacteria. However, establishing and maintaining N2-fixing nodules are expensive for the host plant, relative to taking up N from the soil. Therefore, plants suppress symbiosis when N is plentiful and enhance symbiosis when N is sparse. Here, we show that the nitrate transporter MtNRT2.1 is required for optimal nodule establishment in Medicago truncatula under low-nitrate conditions and the repression of nodulation under high-nitrate conditions. The NIN-like protein (NLP) MtNLP1 is required for MtNRT2.1 expression and regulation of nitrate uptake/transport under low- and high-nitrate conditions. Under low nitrate, the gene encoding the C-terminally encoded peptide (CEP) MtCEP1 was more highly expressed, and the exogenous application of MtCEP1 systemically promoted MtNRT2.1 expression in a compact root architecture 2 (MtCRA2)-dependent manner. The enhancement of nodulation by MtCEP1 and nitrate uptake were both impaired in the Mtnrt2.1 mutant under low nitrate. Our study demonstrates that nitrate uptake by MtNRT2.1 differentially affects nodulation at low- and high-nitrate conditions through the actions of MtCEP1 and MtNLP1.

The Arabidopsis NLP7-HB52/54-VAR2 pathway modulates energy utilization in diverse light and nitrogen conditions

In plants, nitrate is the dominant nitrogen (N)source and a critical nutrient signal regulating various physiological and developmental processes.1,2,3,4 Nitrate-responsive gene regulatory networks are widely believed to control growth, development, and life cycle in addition to N acquisition and utilization,1,2,3,4 and NIN-LIKE PROTEIN (NLP) transcriptional activators have been identified as the master regulators governing the networks.5,6,7 However, it remains to be elucidated how nitrate signaling regulates respective physiological and developmental processes. Here, we have identified a new nitrate-activated transcriptional cascade involved in chloroplast development and the maintenance of chloroplast function in Arabidopsis. This cascade consisting of NLP7 and two homeodomain-leucine zipper (HD-Zip) class I transcription factors, HOMEOBOX PROTEIN52 (HB52) and HB54,8,9 was responsible for nitrate- and light-dependent expression of VAR2 encoding the FtsH2 subunit of the chloroplast FtsH protease involved in the quality control of photodamaged thylakoid membrane proteins.10,11 Consistently, the nitrate-activated NLP7-HB52/54-VAR2 pathway underpinned photosynthetic light energy utilization, especially in high light environments. Furthermore, genetically enhancing the NLP7-HB52/54-VAR2 pathway resulted in improved light energy utilization under high light and low N conditions, a superior agronomic trait. These findings shed light on a new role of nitrate signaling and a novel mechanism for integrating information on N nutrient and light environments, providing a hint for enhancing the light energy utilization of plants in low N environments.