NRT1.1 (NPF6.3/CHL1; AT1G12110) is a plasma-membrane nitrate transporter and nitrate sensor. Proton-coupled nitrate uptake operates over low- and high-affinity regimes regulated by Thr101 phosphorylation, oligomerization and membrane trafficking. Nitrate sensing can be separated genetically from uptake. NPF6.3 also facilitates cellular auxin uptake, particularly under low nitrate, contributing to basipetal auxin movement away from lateral-root tips and restraining their outgrowth. Its nitrate transport and signaling roles influence root-system architecture, guard-cell opening and water loss, nitrate allocation during cadmium stress, and nutrient-dependent flowering responses.
Definition: A signaling process initiated by nitrate perception that changes cellular activity, including nitrate-responsive transcription. The perception and response are distinguished from nitrate uptake and assimilation.
Justification: Retains the earlier review's ontology question with direct uptake/sensing-separation evidence from PMID:19766570. It does not assert that every nitrate-response mechanism follows one obligatory CBL/CIPK chain.
Parent term: response to nitrate
Supporting Evidence:
Definition: Detecting nitrate and converting its recognition into a change that initiates a cellular signaling response, distinguishable from nitrate transmembrane transport.
Justification: The older CHL1 review identified this separate core activity without a molecular-function slot. Uptake/sensing-decoupled mutants directly support the role; the current QuickGO search did not establish a nitrate-specific sensor MF. Use an explicit proposed activity rather than an unrelated transport term.
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005515 protein binding | IPI PMID:19766570 CHL1 functions as a nitrate sensor in plants. | MODIFY | Summary: NPF6.3 binds its regulatory protein kinase CIPK23. Reason: The cited interaction and biochemical phosphorylation identify the partner as a protein kinase, supporting protein kinase binding. This does not assign kinase catalysis to the transporter or infer an independent signaling enzyme activity. Proposed replacements: protein kinase binding Supporting Evidence: file:ARATH/NRT1.1/NRT1.1-uniprot.txt Interacts with CIPK23. PMID:19766570 protein kinase CIPK23 can phosphorylate T101 of CHL1 to maintain a low-level primary response. |
| GO:0005515 protein binding | IPI PMID:31431511 Phosphorylation-Mediated Dynamics of Nitrate Transceptor NRT... | REMOVE | Summary: NRT1.1 phosphorylation-dependent proximity to AtREM1.3 is measured, but bare protein binding is uninformative. Reason: Figure 3 uses FRET-FLIM of fluorescent phosphovariants and remorin in lateral-root cells to assess membrane-microdomain partitioning. Preserve that physical-association evidence; it does not by itself establish remorin as a catalytic cofactor or justify inventing a specific regulatory binding function. Remove only the generic GO:0005515 claim under the project binding policy. Supporting Evidence: PMID:31431511 In plants coexpressing T101D-GFP with AtRem1.3-mCherry, fluorescence lifetime sharply decreased and the FRET efficiency (Ο) increased to 38.7% upon LN treatment and to 18.2% upon HN treatment |
| GO:0005886 plasma membrane | IDA PMID:31431511 Phosphorylation-Mediated Dynamics of Nitrate Transceptor NRT... | ACCEPT | Summary: NPF6.3 occupies the plasma membrane and undergoes regulated internalization. Reason: Native Arabidopsis lateral-root imaging and phosphovariant trafficking experiments directly support plasma-membrane localization. Endosomal signaling after internalization is compatible with this location and does not displace it. Supporting Evidence: PMID:31431511 nonphosphorylatable NRT1.1T101A showed low lateral mobility and oligomerized at the plasma membrane (PM) |
| GO:0006857 oligopeptide transport | IEA GO_REF:0000002 | REMOVE | Summary: The oligopeptide-family mapping incorrectly assigns peptide transport to nitrate-specialized NPF6.3. Reason: IPR018456 identifies a broader transporter lineage with diversified substrates. Direct target binding experiments reject Ala-Ala, and comparison of the nitrate-bound structure with peptide transporters identifies altered substrate-recognition chemistry. The source explicitly explains why AtNRT1.1 cannot recognize or transport peptides; this is target evidence against the propagated substrate, not inference from the NPF family name. Supporting Evidence: PMID:24572366 AtNRT1.1 specifically recognizes both nitrate and chlorate PMID:24572366 does not recognize nitrite, alanine, sulphate, phosphate or the di-peptide Ala-Ala |
| GO:0006950 response to stress | IEA GO_REF:0000117 | MODIFY | Summary: NPF6.3 affects water loss through guard-cell nitrate transport and participates in cadmium-stress nitrate allocation. Reason: The generic electronic stress term can be refined to the experimentally supported water-deprivation response and cadmium-ion response. These are context-specific physiological roles, not evidence that NPF6.3 directly transports cadmium or is a universal stress-protection factor. Proposed replacements: response to water deprivation response to cadmium ion Supporting Evidence: PMID:12509525 chl1 mutants had reduced stomatal opening and reduced transpiration rates in the light or when deprived of CO(2) in the dark. PMID:30972097 NRT1.1 and NRG2 regulated Cd2+ stress-induced NO3 - allocation to roots |
| GO:0009414 response to water deprivation | IMP PMID:12509525 The nitrate transporter AtNRT1.1 (CHL1) functions in stomata... | KEEP AS NON CORE | Summary: Guard-cell nitrate uptake links NPF6.3 to drought susceptibility. Reason: The mutant has less nitrate-dependent stomatal opening and transpiration and consequently greater drought tolerance. Retain a water-deprivation role without reversing the phenotype into a claim that NPF6.3 promotes drought resistance. Supporting Evidence: PMID:12509525 chl1 mutants had reduced stomatal opening and reduced transpiration rates in the light or when deprived of CO(2) in the dark. |
| GO:0010167 response to nitrate | IBA GO_REF:0000033 | ACCEPT | Summary: NPF6.3 contributes directly to nitrate sensing and nitrate-responsive root development. Reason: Nitrate-dependent responses are supported by uptake/sensing separation and nitrate-patch root-colonization experiments. The PAINT ancestral response assignment at PTN000183463 is consistent with this direct target biology; target self-grounding in the source set is legitimate. Supporting Evidence: PMID:19766570 analysis of an uptake- and sensing-decoupled mutant showed that the nitrate transporter CHL1 functions as a nitrate sensor. PMID:17148611 NRT1.1 promotes localized root proliferation independently of any nutritional effect |
| GO:0010167 response to nitrate | IMP PMID:17148611 The Arabidopsis NRT1.1 transporter participates in the signa... | ACCEPT | Summary: NPF6.3 contributes directly to nitrate sensing and nitrate-responsive root development. Reason: Nitrate-dependent responses are supported by uptake/sensing separation and nitrate-patch root-colonization experiments. The PAINT ancestral response assignment at PTN000183463 is consistent with this direct target biology; target self-grounding in the source set is legitimate. Supporting Evidence: PMID:19766570 analysis of an uptake- and sensing-decoupled mutant showed that the nitrate transporter CHL1 functions as a nitrate sensor. PMID:17148611 NRT1.1 promotes localized root proliferation independently of any nutritional effect |
| GO:0010540 basipetal auxin transport | IMP PMID:31431511 Phosphorylation-Mediated Dynamics of Nitrate Transceptor NRT... | ACCEPT | Summary: NPF6.3 contributes to basipetal auxin movement from lateral-root tips under low nitrate. Reason: NRT1.1 performs auxin transport rather than merely causing a downstream developmental phenotype. Full PMID:31431511 Figure 1 measures yeast IAA influx and lateral-root basipetal tracer movement in phosphovariants; PMID:20627075 independently reports nitrate-inhibited cellular auxin uptake. Retain basipetal auxin transport as a core, nitrate-conditioned carrier contribution. This does not convert tissue-level movement out of the root tip into cellular auxin efflux. Supporting Evidence: PMID:20627075 NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. PMID:31431511 These actions could facilitate NRT1.1-dependent auxin flux, thus depleting auxin levels in LRP and inhibiting their outgrowth. PMID:31431511 Basipetal [3H]IAA uptake (counts per minute) in LRs of wild-type, T101A, and T101D seedlings grown for 10 d on medium containing 0.2 and 1 mm of NO3β. |
| GO:0015112 nitrate transmembrane transporter activity | IBA GO_REF:0000033 | ACCEPT | Summary: NPF6.3 catalyzes proton-coupled nitrate uptake over regulated affinity regimes. Reason: Direct oocyte currents and uptake, native mutant uptake defects, and nitrate-bound structures establish nitrate transmembrane transport. PTN000183463 appropriately propagates this experimentally grounded target function; affinity regulation does not require a different substrate-class annotation. Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:24572366 we present both the apo and nitrate-bound crystal structures of Arabidopsis thaliana NRT1.1 |
| GO:0015112 nitrate transmembrane transporter activity | IMP PMID:9844028 The Arabidopsis CHL1 protein plays a major role in high-affi... | ACCEPT | Summary: NPF6.3 catalyzes proton-coupled nitrate uptake over regulated affinity regimes. Reason: Direct oocyte currents and uptake, native mutant uptake defects, and nitrate-bound structures establish nitrate transmembrane transport. PTN000183463 appropriately propagates this experimentally grounded target function; affinity regulation does not require a different substrate-class annotation. The genuine legacy research synthesis provides corroborating context; the primary nitrate uptake and structural experiments remain the decisive evidence. Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:24572366 we present both the apo and nitrate-bound crystal structures of Arabidopsis thaliana NRT1.1 file:ARATH/NRT1.1/NRT1.1-deep-research-falcon.md NPF6.3/CHL1/NRT1.1 is widely regarded as a **dual-affinity** nitrate transporter with **biphasic kinetics**, enabling transport across a wide external nitrate range. |
| GO:0015706 nitrate transmembrane transport | IMP PMID:9844028 The Arabidopsis CHL1 protein plays a major role in high-affi... | ACCEPT | Summary: NPF6.3 performs a nitrate-transport step in plant nitrogen acquisition. Reason: PMID:9844028 identifies high- and low-affinity uptake defects in chl1 mutants; direct heterologous transport independently establishes the carrier step. Retain the process as core, with no claim that NPF6.3 performs subsequent nitrate reduction. Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:9844028 CHL1 is an important component of both the high-affinity and the low-affinity nitrate-uptake systems |
| GO:0016020 membrane | IBA GO_REF:0000033 | MODIFY | Summary: Membrane localization can be refined to the demonstrated plasma membrane. Reason: The general membrane assignment is biologically sound. Target-specific imaging resolves the plasma membrane as a functional location, with regulated trafficking to endosomes; use the already supported precise PM term without treating the inherited membrane claim as false. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN008522918 SUPPORTS TRANSFER The ancestral membrane claim is valid and refined by native NRT1.1 imaging. Proposed replacements: plasma membrane Supporting Evidence: PMID:31431511 nonphosphorylatable NRT1.1T101A showed low lateral mobility and oligomerized at the plasma membrane (PM) |
| GO:0016020 membrane | IEA GO_REF:0000120 | MODIFY | Summary: Membrane localization can be refined to the demonstrated plasma membrane. Reason: The general membrane assignment is biologically sound. Target-specific imaging resolves the plasma membrane as a functional location, with regulated trafficking to endosomes; use the already supported precise PM term without treating the inherited membrane claim as false. Proposed replacements: plasma membrane Supporting Evidence: PMID:31431511 nonphosphorylatable NRT1.1T101A showed low lateral mobility and oligomerized at the plasma membrane (PM) |
| GO:0022857 transmembrane transporter activity | IEA GO_REF:0000002 | MODIFY | Summary: The general transporter activity comprises nitrate transport and conditional auxin influx. Reason: Both substrate activities are directly supported for NPF6.3. Refine the family-level transporter annotation to nitrate transmembrane transporter activity and auxin influx activity; retain both rather than using nitrate specificity to erase the conditional hormone-transport role. Proposed replacements: nitrate transmembrane transporter activity auxin influx transmembrane transporter activity Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:20627075 NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. |
| GO:0048527 lateral root development | IMP PMID:31431511 Phosphorylation-Mediated Dynamics of Nitrate Transceptor NRT... | KEEP AS NON CORE | Summary: Nitrate-dependent NPF6.3 transport and signaling modulate lateral-root development. Reason: NPF6.3 restrains lateral-root outgrowth through auxin redistribution under low nitrate and supports nitrate-responsive proliferation in rich patches. The developmental term is justified, but a universal positive or negative effect would conflate the two nutrient contexts. Supporting Evidence: PMID:20627075 NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. PMID:19766570 analysis of an uptake- and sensing-decoupled mutant showed that the nitrate transporter CHL1 functions as a nitrate sensor. PMID:31431511 These behaviors could promote LR development by suppressing NRT1.1-auxin transport activity on the PM and stimulating Ca2+-ANR1 signaling from the endosome |
| GO:0048573 photoperiodism, flowering | IMP PMID:30972097 NRT1.1 Regulates Nitrate Allocation and Cadmium Tolerance in... | KEEP AS NON CORE | Summary: NPF6.3 contributes to nitrate-dependent photoperiodic flowering, although the original cadmium citation is mismatched. Reason: The complete PMID:30972097 experiments examine cadmium tolerance and nitrate allocation, not flowering. Independent PMID:33643357 compares single and combined transporter mutants and long/short photoperiods, supporting this existing physiological claim. Retain it with the corrected evidence and nutrient context; NPF6.3 is not thereby a photoreceptor or an obligate component of the CO pathway. PMID:31084876 instead supports an FLC-associated route under its distinct conditions. Supporting Evidence: PMID:33643357 NRT1.1 and NRT2.1 were additively involved in LD-induced flowering. PMID:31084876 FLC loss of function could rescue the late flowering phenotype of the chl1-5 mutant |
| GO:0055085 transmembrane transport | IBA GO_REF:0000033 | MODIFY | Summary: The broad transmembrane-transport process can be resolved to nitrate transport and auxin import. Reason: Nitrate transport and auxin cellular uptake are independently measured target activities. These specific processes preserve the ancestral carrier role while avoiding an unsupported assertion that all NPF substrates or directions apply to NPF6.3. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN008522918 SUPPORTS TRANSFER General transmembrane transport is conserved; target assays resolve nitrate and auxin substrates. Proposed replacements: nitrate transmembrane transport auxin import into cell Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:20627075 NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. |
| GO:0055085 transmembrane transport | IEA GO_REF:0000002 | MODIFY | Summary: The broad transmembrane-transport process can be resolved to nitrate transport and auxin import. Reason: Nitrate transport and auxin cellular uptake are independently measured target activities. These specific processes preserve the ancestral carrier role while avoiding an unsupported assertion that all NPF substrates or directions apply to NPF6.3. Proposed replacements: nitrate transmembrane transport auxin import into cell Supporting Evidence: PMID:8453665 Injection of CHL1 mRNA into Xenopus oocytes produces a nitrate- and pH-dependent membrane depolarization, inward current, and nitrate uptake. PMID:20627075 NRT1.1 not only transports nitrate but also facilitates uptake of the phytohormone auxin. |
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Download this section (compressed HTML)Q: Beyond Thr101, how do the CNGC15 Ca2+ channel complex and ABI2/ABA inputs quantitatively tune the balance between CHL1's transport and signaling outputs in vivo?
Q: How do phosphorylation-dependent shifts in monomer/dimer populations, structural flexibility and membrane trafficking jointly determine NRT1.1 affinity and signaling in native cells? The observed populations need not represent a binary all-or-none oligomerization switch.
Q: Can the original PMID:30972097 citation on photoperiodism, flowering be corrected to the intended source? Independent PMID:33643357 now supports a nutrient-conditioned long-day flowering contribution, while PMID:31084876 describes a distinct FLC-associated context.
Experiment: Compare transport-dead but sensing-competent mutants (e.g., Pro492 variants) with sensing-dead variants in planta, measuring both nitrate uptake (15N flux, electrophysiology in oocytes) and primary-nitrate-response gene induction (e.g., NRT2.1, NIA1) to dissect the two functions.
Hypothesis: CHL1's nitrate-sensing (transceptor) function is mechanistically separable from its transport activity and depends on distinct residues.
Type: structure-function and physiological analysis
Experiment: Use phospho-specific antibodies and phosphomimic/non-phosphorylatable Thr101 mutants across a nitrate concentration gradient, combined with cipk23 and cbl1 cbl9 mutants, to quantify how phosphorylation state correlates with apparent Km in vivo.
Hypothesis: The CBL1/CBL9-CIPK23 module phosphorylates Thr101 in a nitrate-concentration- dependent manner to switch transport affinity.
Type: biochemical and genetic analysis
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