NORK (Nodulation receptor kinase; also DMI2, DOES NOT MAKE INFECTIONS 2, and MtSYMRK) is a single-pass transmembrane receptor-like serine/threonine protein kinase of Medicago truncatula with a signal peptide, an extracellular malectin-like domain followed by leucine-rich repeats, a transmembrane helix and a cytoplasmic kinase domain. It is a core component of the common symbiosis signalling pathway (CSSP) shared by the rhizobial root-nodule symbiosis and the arbuscular mycorrhizal (AM) symbiosis: dmi2 null mutants are blocked in Nod factor-induced root hair responses, nodulin gene expression, cortical cell division and nuclear-associated calcium spiking, and fail to form both nodules and AM associations. NORK acts downstream of the LysM receptor kinases (NFP/LYK3 for Nod factors) and upstream of DMI1-dependent calcium spiking and the nuclear decoder CCaMK/DMI3. No ligand for its ectodomain has been identified. The recombinant kinase domain autophosphorylates and trans-phosphorylates substrates in vitro, including the E3 ubiquitin ligase PUB1 (a negative regulator of both symbioses) and the LysM receptor kinase LYK3; it also binds HMG-CoA reductase 1. NORK localises to the plasma membrane of root cells and to the membrane surrounding infection threads in nodules, and is additionally required in nodules for release of rhizobia from infection threads and symbiosome formation. The two symbiotic roles are genetically separable: the kinase-inactive G795E (R38) allele abolishes nodulation but leaves mycorrhization intact.
The preferred accession for this review is Q8L4H4. These relationships are manually assessed.
REFERENCE PROTEOME EQUIVALENT · Reviewed 2026-10-19
Reference proteome: UP000002051
Retain Q8L4H4 as the primary Swiss-Prot accession. The reference sequence is exactly identical across all 925 residues. G7KGF4 is an additional shorter model with 902 identical residues, but A9YWR7 represents the complete primary sequence. The kinase-family search reached its 1000-hit cap; this does not weaken the observed full-length identity but prevents claiming exhaustive paralog retrieval.
MMseqs2 searched 57,093 sequences across UP000002051 at E <= 1e-5 and sensitivity 7.5, returning 1000 hits. Every returned sequence was globally aligned using BLOSUM62 with gap-open -10 and extension -0.5. Scores and sequences for all 1000 returned candidates are retained, with 10 detailed alignment files. The heuristic search does not establish genomic absence or unique locus identity. The search reached its 1000-hit retrieval cap.
file:MEDTR/NORK/NORK-record-evidence.yaml
target_sequence_count: 57093
file:MEDTR/NORK/NORK-record-evidence.yaml
candidate_count: 1000
A9YWR7: 925 identities across 925 global-alignment columns (100.0% identity including gaps), 0 substitutions and 0 gap columns. Query coverage is 100.0%; target coverage is 100.0%. Sequence similarity is interpreted in the manual rationale. Identity over paired residues is 100.0000% (925/925); gap columns are excluded from this denominator.
file:MEDTR/NORK/NORK-record-evidence.yaml
query_length: 925 target_length: 925 alignment_columns: 925 identities: 925 mismatches: 0 gap_columns: 0 identity_percent: 100.0 query_coverage_percent: 100.0 target_coverage_percent: 100.0
The primary accession is outside every searched exact-taxon reference proteome. Current UniProt records place A9YWR7 in UP000002051 and give it the same taxon as the primary.
file:MEDTR/NORK/NORK-record-evidence.yaml
primary_in_any_reference_proteome: false
file:MEDTR/NORK/NORK-record-evidence.yaml
alternative_in_reference_proteome: true
file:MEDTR/NORK/NORK-record-evidence.yaml
same_taxon: true
Sampled UniSave observations: 11-NOV-2015, entry version 79.: no Proteomes cross-reference; 07-OCT-2020, entry version 105.: no Proteomes cross-reference; 27-NOV-2024, entry version 119.: no Proteomes cross-reference. These dated observations do not establish uninterrupted membership/absence, the exact change date, or UniProt's motivation.
file:MEDTR/NORK/NORK-record-evidence.yaml
entry_date: 11-NOV-2015, entry version 79. proteomes: []
file:MEDTR/NORK/NORK-record-evidence.yaml
entry_date: 07-OCT-2020, entry version 105. proteomes: []
file:MEDTR/NORK/NORK-record-evidence.yaml
entry_date: 27-NOV-2024, entry version 119. proteomes: []
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0004672 protein kinase activity | IDA PMID:26839127 PUB1 Interacts with the Receptor Kinase DMI2 and Negatively ... | ACCEPT | Summary: Recombinant DMI2/NORK kinase domain autophosphorylates and directly phosphorylates the E3 ubiquitin ligase PUB1 in vitro (Vernie et al. 2016); independently confirmed by Jayaraman et al. 2017, where NORK trans-phosphorylates casein and kinase-dead LYK3. Reason: Direct biochemical evidence of protein kinase activity on the target itself. The term is general (the more specific serine/threonine kinase terms are also annotated) but correct, and the curator-asserted IDA is consistent with all available data. Supporting Evidence: PMID:26839127 we show that PUB1 also interacts with and is phosphorylated by DOES NOT MAKE INFECTIONS 2, the key symbiotic receptor kinase of the common symbiosis signaling pathway PMID:28267253 the detection of the LYK3 band implies that LYK3 is indeed trans-phosphorylated by NORK |
| GO:0004672 protein kinase activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO mapping from the Ser/Thr kinase catalytic domain and active site signatures. NORK has a canonical kinase domain (residues 596-873) with a predicted catalytic aspartate and ATP-binding lysine. Reason: Electronic inference fully consistent with experimental kinase activity on NORK itself (PMID:26839127, PMID:28267253). Supporting Evidence: PMID:28267253 As expected, the wild-type versions of LYK3 and NORK exhibited auto-phosphorylation (Figure 3A and 3B) and trans-phosphorylation (as indicated by the phosphorylation of casein) activities |
| GO:0004674 protein serine/threonine kinase activity | IEA GO_REF:0000003 | ACCEPT | Summary: EC 2.7.11.1 mapping. UniProt records both serine and threonine phosphorylation (RHEA:17989 and RHEA:46608) from Vernie et al. 2016, and Jayaraman et al. 2017 mapped NORK-phosphorylated serines on LYK3 and NORK autophosphorylation sites including Ser and Thr residues. Reason: Correct and appropriately specific description of the catalytic activity; this is the core molecular function of NORK. Supporting Evidence: PMID:28267253 We identified that sites Ser-269, Ser-273, Ser-307, Ser-323 and Ser-471 of LYK3 (as a control only mLYK3 was used) were phosphorylated by NORK using Mass spectrometry |
| GO:0005515 protein binding | IPI PMID:26839127 PUB1 Interacts with the Receptor Kinase DMI2 and Negatively ... | MODIFY | Summary: Interaction with PUB1 (E4NKF8), a U-box E3 ubiquitin ligase, shown by yeast two-hybrid and co-immunopurification; PUB1 is also a phosphorylation substrate of NORK. Reason: The interaction is real but "protein binding" is uninformative. The cited paper establishes that the partner is an E3 ubiquitin ligase, so the more informative "ubiquitin protein ligase binding" (GO:0031625) is directly supported by the same evidence. The functional relationship is kinase-substrate (NORK phosphorylates PUB1); PUB1 did not detectably ubiquitinate DMI2 according to the deep-research summary of the full text. Proposed replacements: ubiquitin protein ligase binding Supporting Evidence: PMID:26839127 PUB1, an E3 ubiquitin ligase, which interacts with and is phosphorylated by the LYK3 symbiotic receptor kinase PMID:26839127 we show that PUB1 also interacts with and is phosphorylated by DOES NOT MAKE INFECTIONS 2 file:MEDTR/NORK/NORK-deep-research-falcon.md Yeast two-hybrid and co-immunopurification support DMI2–PUB1 binding; recombinant DMI2 autophosphorylates and directly phosphorylates PUB1 in vitro |
| GO:0005524 ATP binding | IEA GO_REF:0000002 | ACCEPT | Summary: Kinase domain ATP-binding signature (residues 602-610 and Lys624). ATP binding is intrinsic to the demonstrated kinase activity. Reason: Correct structural inference for an active protein kinase; non-informative beyond the kinase activity terms but not wrong. Supporting Evidence: PMID:28267253 As expected, the wild-type versions of LYK3 and NORK exhibited auto-phosphorylation |
| GO:0016020 membrane | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping (single-pass membrane protein, by similarity/topology). NORK has a signal peptide and a single transmembrane helix (522-542). Reason: Correct but general. Experimental localisation shows NORK is at the plasma membrane and the infection-thread membrane (PMID:16006515); a NEW plasma membrane annotation is proposed below to capture the specific location. Supporting Evidence: PMID:16006515 The protein locates to the host cell plasma membrane and to the membrane surrounding the infection threads. |
| GO:0046777 protein autophosphorylation | IDA PMID:26839127 PUB1 Interacts with the Receptor Kinase DMI2 and Negatively ... | ACCEPT | Summary: Purified DMI2 kinase domain autophosphorylates in vitro (Vernie et al. 2016); autophosphorylation was also shown by Jayaraman et al. 2017, who mapped at least 37 NORK phosphosites and showed the G794E (R38) mutant lacks autophosphorylation. Reason: Directly demonstrated on the target. Autophosphorylation is a regulatory property of the kinase (SYMRK kinase activity depends on its phosphorylation status) rather than its biological output, but the annotation is accurate. Supporting Evidence: file:MEDTR/NORK/NORK-deep-research-falcon.md Purified NORK/DMI2 intracellular kinase autophosphorylates and trans-phosphorylates substrates; kinase-dead G794E lacks activity PMID:28267253 At least 37 phosphorylation sites of NORK were identified in this study |
| GO:0106310 protein serine kinase activity | EXP PMID:26839127 PUB1 Interacts with the Receptor Kinase DMI2 and Negatively ... | ACCEPT | Summary: UniProt Rhea-based curation (RHEA:17989) from the PUB1 trans-phosphorylation and autophosphorylation assays of Vernie et al. 2016. Serine phosphorylation of a substrate by NORK is independently confirmed by MS-mapped serines on LYK3 (Jayaraman et al. 2017). Reason: Supported by direct biochemical data; consistent with the serine/threonine kinase classification. Supporting Evidence: PMID:26839127 PUB1 also interacts with and is phosphorylated by DOES NOT MAKE INFECTIONS 2 PMID:28267253 We identified that sites Ser-269, Ser-273, Ser-307, Ser-323 and Ser-471 of LYK3 (as a control only mLYK3 was used) were phosphorylated by NORK |
| GO:0106310 protein serine kinase activity | IEA GO_REF:0000116 | ACCEPT | Summary: Electronic mapping from the UniProt Rhea catalytic-activity annotation, itself experimentally based (PMID:26839127). Reason: Consistent with experimental evidence for serine phosphorylation by NORK. Supporting Evidence: PMID:28267253 We identified that sites Ser-269, Ser-273, Ser-307, Ser-323 and Ser-471 of LYK3 (as a control only mLYK3 was used) were phosphorylated by NORK |
| GO:0009877 nodulation | IMP PMID:11006338 Four genes of Medicago truncatula controlling components of ... | NEW | Summary: dmi2 mutants are Nod- and blocked in all early Nod factor responses (root hair deformation, nodulin expression, cortical cell division) and in Nod factor-induced calcium spiking; reduced DMI2 expression in nodules blocks release of rhizobia from infection threads and symbiosome formation. NORK is the receptor-kinase step of the Nod factor signal transduction pathway. Reason: Nodulation is the best-established biological role of NORK and is absent from GOA. Participation test - NORK is not a substrate or passive requirement; it is an active signal-transducing kinase in the Nod factor signalling cascade that phosphorylates pathway components (LYK3, PUB1) and whose kinase activity is specifically required for nodulation (the kinase-dead G795E/R38 allele is Nod- but Myc+). Comparator check - the other Medicago signalling components in the same pathway carry this term: LYK3 (Q6UD73, IMP PMID:12947035), CCaMK/DMI3 (Q6RET7, IMP PMID:22874912) and NFP. Its absence from NORK is a curation gap rather than a convention. Supporting Evidence: PMID:11006338 Mutants in four genes ( DMI1 , DMI2 , DMI3 , and NSP ) were pleiotropically affected in Nod factor responses, indicating that these genes are required for a Nod factor–activated signal transduction pathway that leads to symbiotic responses such as root hair deformations, expressions of nodulin genes, and cortical cell divisions. PMID:11078514 We defined two genes, DMI1 and DMI2, required in common for early steps of infection and nodulation and for calcium spiking. PMID:16006515 At least one of these genes, encoding the receptor kinase DOES NOT MAKE INFECTIONS 2 (DMI2) is essential for symbiosome formation. PMID:28267253 had no effect on mycorrhization of plants but impaired the ability of plants to form nodules |
| GO:0036377 arbuscular mycorrhizal association | IMP PMID:11006338 Four genes of Medicago truncatula controlling components of ... | NEW | Summary: dmi2 mutants of M. truncatula cannot establish AM symbiosis; NORK/SYMRK is the receptor-kinase entry point of the common symbiosis signalling pathway used by both rhizobia and AM fungi. Reason: The AM role is equally well established genetically and is missing from GOA. Kept as a distinct process term from nodulation because the two are genetically separable (G795E/R38 abolishes nodulation without affecting mycorrhization), so NORK's AM function does not rely on the same kinase requirement. Participation - NORK is a signalling component of the CSSP that transduces the symbiotic signal towards nuclear calcium spiking. Comparator - rice CCaMK, a downstream CSSP component, carries this term in this repository's review (genes/ORYSJ/CCAMK). Direct binding of a Myc factor to NORK is not established, so no receptor/ligand-binding term is proposed. Supporting Evidence: PMID:11006338 dmi1 , dmi2 , and dmi3 mutants are also unable to establish a symbiotic association with endomycorrhizal fungi, indicating that there are at least three common steps to nodulation and endomycorrhization in M. truncatula PMID:28267253 Also, NORK is essential for the formation of a symbiotic association with arbuscular mycorrhizal fungi PMID:26839127 the key symbiotic receptor kinase of the common symbiosis signaling pathway, required for both the rhizobial and the arbuscular mycorrhizal (AM) endosymbioses |
| GO:0005886 plasma membrane | IDA PMID:16006515 Formation of organelle-like N2-fixing symbiosomes in legume ... | NEW | Summary: Immunolocalisation/fusion analysis in M. truncatula nodules places DMI2 at the host cell plasma membrane and the infection-thread membrane. Reason: More specific, experimentally supported location than the existing IEA "membrane" annotation; consistent with a single-pass receptor-like kinase. The comparator LYK3 carries plasma membrane by IDA. Supporting Evidence: PMID:16006515 The protein locates to the host cell plasma membrane and to the membrane surrounding the infection threads. |
Loading supporting content…
Download this section (compressed HTML)Q: What is the ligand (if any) of the NORK malectin-like/LRR ectodomain, and is ectodomain cleavage/release of the malectin-like domain a regulated step in rhizobial and AM signalling?
Q: Why is NORK kinase activity required for nodulation but apparently dispensable for AM colonisation (G795E/R38 is Nod- Myc+)? Does NORK act as a scaffold for AM signalling, or does residual activity suffice?
Q: Is the in vitro phosphorylation of LYK3 and PUB1 by NORK relevant in vivo, and what links NORK at the plasma membrane to DMI1-dependent nuclear calcium spiking (e.g. HMGR1-derived mevalonate)?
Experiment: Complement M. truncatula dmi2 null mutants with kinase-dead and phosphosite-mutant NORK variants and score nodulation, AM colonisation and nuclear calcium spiking separately.
Hypothesis: NORK kinase activity is required for the nodulation branch but not for AM signalling, so the two symbiotic outputs depend on different NORK activities.
Type: genetic complementation with structure-function mutants
Experiment: Phosphoproteomics of roots expressing wild-type versus kinase-dead NORK after Nod factor or Myc-LCO treatment to identify in vivo substrates.
Hypothesis: LYK3 and PUB1 are phosphorylated in a NORK-dependent manner in planta.
Type: quantitative phosphoproteomics
Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)