CNNM4

UniProt ID: A0A8I3PI07
Organism: Canis lupus familiaris
Review Status: DRAFT
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Gene Description

CNNM4 (cyclin and CBS domain divalent metal cation transport mediator 4) is a multi-pass plasma-membrane protein of the CNNM/ACDP family. The dog protein (796 aa) has a signal peptide, an N-terminal extracellular immunoglobulin-like domain, the four-helix CNNM transmembrane domain, a cytoplasmic pair of CBS domains that binds MgATP, and a C-terminal cyclic nucleotide-binding homology domain. In mammals CNNM4 sits at the basolateral membrane of intestinal epithelial cells and enamel-forming ameloblasts, where it extrudes Mg2+ from the cell in exchange for extracellular Na+. This basolateral efflux is the exit step of transcellular magnesium absorption, so CNNM4 is needed for systemic magnesium balance and for enamel formation. MgATP binding to the CBS pair organises the full-length protein into a dimer-of-dimers assembly, and CNNM4 also associates with the TRPM7 channel and stimulates divalent-cation entry through it. In humans, biallelic loss-of-function variants cause Jalili syndrome (cone-rod dystrophy with amelogenesis imperfecta). No functional study of the dog protein itself has been reported; its function is inferred from its placement in the CNNM4 subfamily and 93% identity to human CNNM4.

Existing Annotations Review

GO Term Evidence Action Reason
GO:0005886 plasma membrane
IBA
GO_REF:0000033 		ACCEPT
Summary: PAINT node PTN000248898 assigns plasma-membrane activity across the CNNM family, seeded by experimental annotations of fly uex, mouse Cnnm1/2/4, human CNNM2/CNNM4 and worm homologs. The dog protein is inside this clade (PTHR12064:SF26) and has the signal peptide and four predicted transmembrane helices of the family.
Reason: The target is inside the clade that inherited the location, and nothing in its sequence suggests divergence (the transmembrane domain is intact; 93% identity to human CNNM4). Mammalian CNNM4 extrudes Mg2+ across the plasma membrane and localizes to the basolateral surface of epithelia.
Supporting Evidence:
GO_REF:0000033
PMID:24339795
Endogenous CNNM4 is highly expressed in the mature ameloblasts and intestinal epithelia, and localizes at their basolateral membrane.
file:CANLF/A0A8I3PI07/A0A8I3PI07-bioinformatics/RESULTS.md
Dog vs human CNNM4: identity 0.926 over 775 aligned columns (similarity 0.961).
GO:0010960 magnesium ion homeostasis
IBA
GO_REF:0000033 		ACCEPT
Summary: Eukaryote-level PAINT node PTN008532859 (mouse Cnnm4, human CNNM2, worm and zebrafish CNNMs). CNNM4-knockout mice develop hypomagnesemia from defective intestinal Mg2+ absorption.
Reason: Magnesium homeostasis is a core, family-wide process and the dog protein is a CNNM4-subfamily member with all functional domains. CNNM4 directly performs the basolateral Mg2+ efflux step, so this is participation and not only necessity.
Supporting Evidence:
GO_REF:0000033
PMID:24339795
In this study, we generated CNNM4-knockout mice; these mice showed defects in amelogenesis and intestinal Mg2+ absorption.
GO:0015081 sodium ion transmembrane transporter activity
IBA
GO_REF:0000033 		MODIFY
Summary: PAINT node PTN002635769 (Euteleostomi, seeded by mouse Cnnm4) propagates sodium transporter activity together with magnesium transporter activity. In CNNM4-expressing cells, Mg2+ extrusion requires extracellular Na+ and is accompanied by Na+ entry, and the human CNNM4 structure is described as promoting Mg2+/Na+ exchange.
Reason: The sodium movement is real but is the counter-ion arm of a coupled Mg2+/Na+ exchange, not independent sodium transport. The more precise coupled term, GO:0061768 magnesium:sodium antiporter activity, captures both IBA MF rows from this node and matches the human CNNM4 review. The node is CNNM4-specific and the target is in the same subfamily (SF26), so transfer of the coupled activity is warranted.
Propagation Review
Root cause: TERM SCOPING PROBLEM
Failure modes: GRANULARITY MISMATCH
Sources checked:
PANTHER:PTN002635769 SUPPORTS TRANSFER
Euteleostomi CNNM4-subfamily node; the dog target (PTHR12064:SF26) descends from it. The function transfers; only the term granularity is refined.
MGI:MGI:2151060 SUPPORTS TRANSFER
Mouse Cnnm4, the sole seed; experimental Mg2+ extrusion with Na+ dependence.
Supporting Evidence:
GO_REF:0000033
PMID:24339795
In the absence of extracellular Na+, Mg2+ depletion did not induce Mg2+ extrusion (Phase 3), but restoration of Na+ instantaneously caused Mg2+ extrusion (Phase 4).
PMID:42497866
the outward-facing state, thereby promoting Mg2+/Na+ exchange.
GO:0015095 magnesium ion transmembrane transporter activity
IBA
GO_REF:0000033 		MODIFY
Summary: Same CNNM4-specific PAINT node (PTN002635769, seeded by mouse Cnnm4). CNNM4 extrudes Mg2+ from cells in a Na+-dependent manner.
Reason: Correct but less specific than the evidence supports: CNNM4 moves Mg2+ out of the cell in exchange for Na+. GO:0061768 magnesium:sodium antiporter activity is the specific descendant and is the replacement used for the sodium-transporter row as well.
Propagation Review
Root cause: TERM SCOPING PROBLEM
Failure modes: GRANULARITY MISMATCH
Sources checked:
PANTHER:PTN002635769 SUPPORTS TRANSFER
Euteleostomi CNNM4-subfamily node; the dog target (PTHR12064:SF26) descends from it. The function transfers; only the term granularity is refined.
MGI:MGI:2151060 SUPPORTS TRANSFER
Mouse Cnnm4, the sole seed; experimental Mg2+ extrusion with Na+ dependence.
Supporting Evidence:
GO_REF:0000033
PMID:24339795
As shown in Figure 4D, Mg2+ depletion from the extracellular medium induced not only the decrease of intracellular Mg2+ but also the increase of intracellular Na+.
GO:0015693 magnesium ion transport
IBA
GO_REF:0000033 		ACCEPT
Summary: CNNM4-specific PAINT node PTN002635769. CNNM4 mediates basolateral Mg2+ extrusion in transcellular epithelial Mg2+ transport.
Reason: Correct and inherited; the gene also carries GO:1903830 magnesium ion transmembrane transport, which is the more specific process, so this broad term is consistent with it rather than a competing claim.
Supporting Evidence:
GO_REF:0000033
PMID:24339795
Functional analyses at the molecular and organismal levels revealed a common role for CNNM4 in mediating transcellular Mg2+ transport by basolateral Mg2+ extrusion.
GO:0005886 plasma membrane
IEA
GO_REF:0000120 		ACCEPT
Summary: Combined electronic annotation (human CNNM4 ortholog, UniProt SubCell mapping, PANTHER). Agrees with the IBA row for the same term.
Reason: Consistent with the IBA plasma-membrane row and with mammalian CNNM4 localization; the more specific basolateral location is annotated separately for polarized epithelia.
Supporting Evidence:
PMID:24339795
Endogenous CNNM4 is highly expressed in the mature ameloblasts and intestinal epithelia, and localizes at their basolateral membrane.
GO:0010960 magnesium ion homeostasis
IEA
GO_REF:0000120 		ACCEPT
Summary: InterPro ACDP family (IPR045095) and PANTHER mapping; same term as the IBA row.
Reason: Same claim as the accepted IBA row; supported by mouse CNNM4 knockout physiology.
Supporting Evidence:
PMID:24339795
In this study, we generated CNNM4-knockout mice; these mice showed defects in amelogenesis and intestinal Mg2+ absorption.
GO:0015693 magnesium ion transport
IEA
GO_REF:0000118 		ACCEPT
Summary: TreeGrafter assignment from the PANTHER CNNM4 subfamily; same term as the IBA row.
Reason: Same claim as the accepted IBA row.
Supporting Evidence:
PMID:24339795
Functional analyses at the molecular and organismal levels revealed a common role for CNNM4 in mediating transcellular Mg2+ transport by basolateral Mg2+ extrusion.
GO:0016020 membrane
IEA
GO_REF:0000104 		MODIFY
Summary: UniRule assignment of the generic membrane term. True (four predicted transmembrane helices) but much less specific than the plasma-membrane location the gene already carries.
Reason: Correct but general. The specific supported location, GO:0005886 plasma membrane, is already annotated to this gene by IBA and IEA.
Proposed replacements: plasma membrane
Supporting Evidence:
PMID:24339795
Endogenous CNNM4 is highly expressed in the mature ameloblasts and intestinal epithelia, and localizes at their basolateral membrane.
GO:0022857 transmembrane transporter activity
IEA
GO_REF:0000120 		MODIFY
Summary: PANTHER/UniRule assignment of the generic transporter term (PAINT node PTN000248941 lineage). True but non-specific.
Reason: Correct but general; the specific supported activity is Mg2+/Na+ exchange (GO:0061768 magnesium:sodium antiporter activity), the same replacement proposed for the IBA transporter rows.
Supporting Evidence:
PMID:42497866
the outward-facing state, thereby promoting Mg2+/Na+ exchange.
GO:0035725 sodium ion transmembrane transport
IEA
GO_REF:0000108 		ACCEPT
Summary: Logical inference from the sodium transporter activity row. CNNM4-expressing cells take up Na+ while extruding Mg2+.
Reason: Na+ entry is the counter-ion arm of CNNM4 Mg2+/Na+ exchange, performed by CNNM4 itself, so the process holds even though the MF is refined to the antiporter term.
Supporting Evidence:
PMID:24339795
As shown in Figure 4A, HEK293 cells transfected with CNNM4-FLAG contained more sodium and less magnesium in comparison to control vector-transfected cells, consistent with the occurrence of Mg2+ extrusion.
GO:0055085 transmembrane transport
IEA
GO_REF:0000108 		MODIFY
Summary: Logical inference from the generic transporter activity row. True but non-specific.
Reason: Correct but general. The specific supported processes, GO:1903830 magnesium ion transmembrane transport and GO:0035725 sodium ion transmembrane transport, are both already annotated to this gene.
Supporting Evidence:
PMID:24339795
As shown in Figure 4D, Mg2+ depletion from the extracellular medium induced not only the decrease of intracellular Mg2+ but also the increase of intracellular Na+.
GO:1903830 magnesium ion transmembrane transport
IEA
GO_REF:0000108 		ACCEPT
Summary: Logical inference from magnesium transporter activity. CNNM4 extrudes Mg2+ across the basolateral plasma membrane.
Reason: The core process carried out by CNNM4; supported by mammalian efflux assays.
Supporting Evidence:
PMID:24339795
Functional analyses at the molecular and organismal levels revealed a common role for CNNM4 in mediating transcellular Mg2+ transport by basolateral Mg2+ extrusion.
GO:0015081 sodium ion transmembrane transporter activity
IEA
GO_REF:0000120 		MODIFY
Summary: Combined electronic transfer from mouse Cnnm4 (Q69ZF7) and PANTHER; same claim as the IBA row for this term.
Reason: Same refinement as the IBA row; Na+ movement is coupled to Mg2+ efflux.
Supporting Evidence:
PMID:24339795
In the absence of extracellular Na+, Mg2+ depletion did not induce Mg2+ extrusion (Phase 3), but restoration of Na+ instantaneously caused Mg2+ extrusion (Phase 4).
GO:0015095 magnesium ion transmembrane transporter activity
IEA
GO_REF:0000120 		MODIFY
Summary: Combined electronic transfer from mouse Cnnm4 (Q69ZF7) and PANTHER; same claim as the IBA row for this term.
Reason: Same refinement as the IBA row.
Supporting Evidence:
PMID:42497866
the outward-facing state, thereby promoting Mg2+/Na+ exchange.
GO:0016323 basolateral plasma membrane
IEA
GO_REF:0000107 		ACCEPT
Summary: Ensembl Compara transfer from mouse Cnnm4 (Q69ZF7). Endogenous mouse CNNM4 is at the basolateral membrane of intestinal epithelia and ameloblasts, and human CNNM4 expressed in polarized MDCK (dog kidney) cells is also basolateral.
Reason: Ortholog transfer within the CNNM4 subfamily of a location that underlies the vectorial transport function. This is the location in polarized epithelia; it does not exclude plasma-membrane localization in non-polarized cells.
Supporting Evidence:
PMID:24339795
Endogenous CNNM4 is highly expressed in the mature ameloblasts and intestinal epithelia, and localizes at their basolateral membrane.
GO:0030003 intracellular monoatomic cation homeostasis
IEA
GO_REF:0000107 		MODIFY
Summary: Ensembl Compara transfer from human CNNM4. The human source annotation rests on metal-toxicity assays; the cation CNNM4 is known to handle is Mg2+, and CNNM4 efflux directly lowers intracellular Mg2+.
Reason: Correct but general. CNNM4-dependent efflux lowers intracellular Mg2+, so the specific descendant GO:0010961 intracellular magnesium ion homeostasis is supported (it is_a GO:0030003 and GO:0010960, the latter already annotated). Other cations are not established as CNNM4 substrates.
Supporting Evidence:
PMID:24339795
As shown in Figure 4D, Mg2+ depletion from the extracellular medium induced not only the decrease of intracellular Mg2+ but also the increase of intracellular Na+.
GO:0032991 protein-containing complex
IEA
GO_REF:0000107 		KEEP AS NON CORE
Summary: Ensembl Compara transfer from human CNNM4. Full-length human CNNM4 forms an MgATP-dependent asymmetric tetramer, and CNNM4 associates with TRPM7.
Reason: True (homo-oligomer; TRPM7 association) but no specific GO complex term exists for either assembly, so the broad term is as precise as available. Kept as non-core.
Supporting Evidence:
PMID:42497866
These structures reveal that CNNM4 does not assemble as a simple symmetric dimer but forms an asymmetric tetramer organized as a dimer-of-dimers.

Core Functions

CNNM4 exchanges intracellular Mg2+ for extracellular Na+ across the basolateral plasma membrane of polarized epithelia (intestine, ameloblasts), providing the exit step of transcellular magnesium absorption and supporting systemic and intracellular Mg2+ balance. MgATP binding to the CBS pair organises the full-length protein into an asymmetric dimer-of-dimers. For the dog protein this is inferred from CNNM4-subfamily placement (PTHR12064:SF26, PAINT node PTN002635769) and 93% identity to human CNNM4.

Molecular Function:
magnesium:sodium antiporter activity
Directly Involved In:
magnesium ion transmembrane transport sodium ion transmembrane transport magnesium ion homeostasis
Cellular Locations:
basolateral plasma membrane
Supporting Evidence:
  • PMID:24339795
    In the absence of extracellular Na+, Mg2+ depletion did not induce Mg2+ extrusion (Phase 3), but restoration of Na+ instantaneously caused Mg2+ extrusion (Phase 4).
  • PMID:42497866
    the outward-facing state, thereby promoting Mg2+/Na+ exchange.
  • file:CANLF/A0A8I3PI07/A0A8I3PI07-bioinformatics/RESULTS.md
    Dog vs human CNNM4: identity 0.926 over 775 aligned columns (similarity 0.961).

References

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Suggested Questions for Experts

Q: GO:0061768 magnesium:sodium antiporter activity is written as Na+(in) + Mg2+(out) = Na+(out) + Mg2+(in), whereas CNNM4 extrudes Mg2+ and takes up Na+. Should the definition be direction-neutral, or is a separate export-direction term needed?

Q: Does the ~21-residue C-terminal insertion in the dog CNNM4 gene model correspond to an alternative exon present in other carnivores, and does it affect the CNBH domain?

Q: Should the TRPM7-stimulating (ion channel regulator) activity shown for human CNNM4 be propagated phylogenetically to CNNM4 orthologs?

External Prediction Reviews

These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.

ProtNLM2 External predictions

View prediction review YAML · A0A8I3PI07-protnlm-predictions-review.yaml · Review status: COMPLETE

ProtNLM2 emitted three GO predictions for dog CNNM4, all broad terms that are correct for a CNNM4-subfamily Mg2+/Na+ exchanger at the basolateral plasma membrane. Each is less specific than existing supported annotations (basolateral plasma membrane, magnesium ion transmembrane transporter activity, magnesium ion transport), so all three are LSP.

Source documents: genes/CANLF/A0A8I3PI07/A0A8I3PI07-uniprot.txt · genes/CANLF/A0A8I3PI07/A0A8I3PI07-ai-review.yaml · publications/PMID_24339795.md · publications/PMID_25449265.md · publications/PMID_42497866.md

Review score: 2 = concordant with evidence; 1 = uncertain; 0 = discordant with evidence. This is an assessment score, not a model probability.

GO:0005886 plasma membrane GO_CC
LSP — Less precise than existing annotation Review score: 2/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
Review rationale: The target is placed in the CNNM4 subfamily (PANTHER PTHR12064:SF26) and has a signal peptide, four predicted transmembrane helices within the CNNM transmembrane domain, and a cytoplasmic CBS domain, the architecture of a multi-pass plasma-membrane transporter. Mouse CNNM4 localizes to the basolateral membrane of intestinal epithelia and ameloblasts (PMID:24339795), and endogenous CNNM4 in Madin-Darby canine kidney cells is basolateral (PMID:25449265), which is direct evidence in the dog. Plasma membrane is already annotated to the target, and the more specific basolateral plasma membrane (GO:0016323, is_a descendant of GO:0005886 per OLS4) is an existing annotation that the gene review accepts. The prediction is correct but less specific than that supported annotation.
Supporting Evidence:
  • file:CANLF/A0A8I3PI07/A0A8I3PI07-uniprot.txt: "DR PANTHER; PTHR12064:SF26; METAL TRANSPORTER CNNM4; 1. ... FT SIGNAL 1..37 ... FT TRANSMEM 182..206 ... FT /note="CNNM transmembrane" ... FT /note="CBS""
  • PMID:24339795: "CNNM4 localizes to the basolateral membrane"
  • PMID:25449265: "We first confirmed the basolateral localization of both endogenous and ectopically expressed CNNM4 in Madin-Darby Canine Kidney cells"
GO:0022857 transmembrane transporter activity GO_MF
LSP — Less precise than existing annotation Review score: 2/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
Review rationale: The target belongs to the CNNM4 subfamily (PTHR12064:SF26) and carries the CNNM transmembrane and CBS domains. Mouse CNNM4 extrudes Mg2+ from cells in a Na+-dependent manner, mediating transcellular Mg2+ transport (PMID:24339795), and structural work on human CNNM4 describes Mg2+/Na+ exchange (PMID:42497866); the dog protein is in the same subfamily, so the activity transfers by phylogenetic inference rather than direct observation. Generic transmembrane transporter activity is true but uninformative: the target already carries magnesium ion transmembrane transporter activity (GO:0015095, a descendant of GO:0022857 per OLS4) and sodium ion transmembrane transporter activity, and the gene review refines these to magnesium:sodium antiporter activity. The prediction is therefore less precise than existing supported annotations.
Supporting Evidence:
  • file:CANLF/A0A8I3PI07/A0A8I3PI07-uniprot.txt: "DR PANTHER; PTHR12064:SF26; METAL TRANSPORTER CNNM4; 1. ... FT SIGNAL 1..37 ... FT TRANSMEM 182..206 ... FT /note="CNNM transmembrane" ... FT /note="CBS""
  • PMID:24339795: "Functional analyses at the molecular and organismal levels revealed a common role for CNNM4 in mediating transcellular Mg2+ transport by basolateral Mg2+ extrusion."
  • PMID:24339795: "In the absence of extracellular Na+, Mg2+ depletion did not induce Mg2+ extrusion (Phase 3), but restoration of Na+ instantaneously caused Mg2+ extrusion (Phase 4)."
  • PMID:42497866: "the outward-facing state, thereby promoting Mg2+/Na+ exchange."
GO:0006811 monoatomic ion transport GO_BP
LSP — Less precise than existing annotation Review score: 2/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
Review rationale: CNNM4 mediates basolateral Mg2+ extrusion coupled to Na+ in mouse epithelia (PMID:24339795), and the dog target inherits this function through its CNNM4 subfamily placement (PTHR12064:SF26) and conserved transmembrane and CBS domain architecture. Monoatomic ion transport is not itself annotated, but the target already carries magnesium ion transport (GO:0015693), magnesium ion transmembrane transport and sodium ion transmembrane transport, which the gene review accepts; OLS4 confirms GO:0015693 is a descendant of GO:0006811 via metal ion and monoatomic cation transport. The prediction is correct but discards the established ion specificity, so it is less precise than existing annotations.
Supporting Evidence:
  • file:CANLF/A0A8I3PI07/A0A8I3PI07-uniprot.txt: "DR PANTHER; PTHR12064:SF26; METAL TRANSPORTER CNNM4; 1. ... FT SIGNAL 1..37 ... FT TRANSMEM 182..206 ... FT /note="CNNM transmembrane" ... FT /note="CBS""
  • PMID:24339795: "Functional analyses at the molecular and organismal levels revealed a common role for CNNM4 in mediating transcellular Mg2+ transport by basolateral Mg2+ extrusion."
  • PMID:24339795: "In the absence of extracellular Na+, Mg2+ depletion did not induce Mg2+ extrusion (Phase 3), but restoration of Na+ instantaneously caused Mg2+ extrusion (Phase 4)."

Deep Research

Falcon

(A0A8I3PI07-deep-research-falcon.md)

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📚 Additional Documentation

Notes

(A0A8I3PI07-notes.md)

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Bioinformatics Results

(RESULTS.md)

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