cmc4 (systematic name SPAC4F10.22; synonym tam2) is a small (70-residue) mitochondrial intermembrane space (IMS) protein of the twin CX9C / coiled-coil- helix-coiled-coil-helix (CHCH) family, the sole fission-yeast member of the conserved CMC4 (Cx9C motif-containing protein 4) family (PANTHER PTHR15590, Pfam PF08991, InterPro IPR027179). Its two CX9C motifs form a helical hairpin stabilized by two intramolecular disulfide bonds; like other twin CX9C proteins it is imported into the IMS and oxidatively folded by the MIA40 (Mia40/CHCHD4)- Erv1 disulfide relay, which recognizes the cysteine motifs. CMC4 orthologs are broadly conserved from yeasts to human (human CMC4, P56277). The family is associated with cytochrome c oxidase (COX) biogenesis: in human cells CMC4 physically associates with the COX assembly factor SCO1, implicating it in COX II sub-assembly, although CMC4 is non-essential for respiration and no specific molecular activity has been demonstrated for it. In S. pombe the gene was identified as a meiotically regulated transcript (tam2), and its deletion is viable with no reported growth or respiratory phenotype.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005758 mitochondrial intermembrane space | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic (IBA) inference of mitochondrial intermembrane space localization from the CMC4 family (PANTHER PTHR15590 / PTN000400520). This is well supported: the twin CX9C / CHCH fold and the MIA40-Erv1 import route are diagnostic of soluble IMS proteins, and the human/yeast orthologs are IMS-localized. This is the single substantive, defensible annotation for cmc4 and represents its core cellular context. Reason: Localization is strongly supported by family membership (twin CX9C, MIA40-import) and ortholog data; the IBA inference from PTHR15590 is appropriate. Note the localization has not been demonstrated experimentally in S. pombe itself. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000400520 Β· CMC4 family (PTHR15590) SUPPORTS TRANSFER Twin CX9C / MIA40-imported IMS protein family; human and yeast orthologs are IMS-localized. Supporting Evidence: PMID:33498264 CMC4 is a CX9C family member that localizes to the mitochondrial IMS |
| GO:0005758 mitochondrial intermembrane space | IEA GO_REF:0000044 | ACCEPT | Summary: Electronic annotation of the same localization derived from the UniProtKB/Swiss-Prot subcellular-location keyword (SL-0169). Redundant with, and consistent with, the IBA annotation above. Reason: Consistent with the family-supported IMS localization; the electronic SubCell mapping mirrors the UniProt annotation and does not overreach. |
| GO:0003674 molecular_function | ND GO_REF:0000015 | ACCEPT | Summary: Molecular function is recorded as ND (no data). No specific biochemical activity has been demonstrated for cmc4 or any CMC4 ortholog: unlike the related copper chaperones Cmc1/Cmc2, CMC4 has no demonstrated metal binding or catalytic activity, and its only functional lead is a physical association with SCO1 in human cells. The ND root placeholder honestly reflects a genuine molecular-function knowledge gap. Reason: No experimental or defensible computational molecular-function term can be asserted. Retaining the ND annotation correctly signals an undetermined molecular function rather than forcing an unsupported term. Knowledge gap: The molecular activity of cmc4 is undetermined. It is unknown whether it acts as a metallochaperone, a redox/disulfide-relay accessory factor, or a protein-interaction adaptor within the mitochondrial IMS, and no direct binding partner has been identified in S. pombe. OPEN BIOLOGY MF_DARK Resolve: Affinity/proximity proteomics of tagged cmc4 in S. pombe to identify direct partners; metal-binding and thiol-redox assays on recombinant protein to test metallochaperone vs disulfide-relay-accessory models. "determined that CMC4 interacts with SCO1 in human cell cultures" β PMID:33498264 |
| GO:0008150 biological_process | ND GO_REF:0000015 | ACCEPT | Summary: Biological process is recorded as ND (no data). A COX II sub-assembly role is only inferred from the human SCO1 interaction, is non-essential, and has not been tested in S. pombe; tam2Ξ is viable with no reported respiratory or meiotic phenotype despite the transcript being meiotically regulated. The ND placeholder honestly reflects an undetermined biological process. Reason: No experimentally supported biological process for cmc4 in S. pombe. The COX-assembly link is cross-species and non-essential; the meiotic transcript regulation (tam2) has no assigned meiotic function. ND is the correct, honest annotation. Knowledge gap: The in vivo biological role of cmc4 in S. pombe is unknown: whether it contributes to cytochrome c oxidase assembly/respiration (as suggested for the human ortholog) or has a distinct meiosis-related role implied by its tam2 expression pattern is undetermined, and no deletion phenotype has been reported. OPEN BIOLOGY BP_DARK Resolve: Phenotyping of cmc4Ξ (respiratory growth on non-fermentable carbon, COX activity/assembly; sporulation, spore viability and meiotic timing) and epistasis with cox11/sco1 orthologs. "While tam2 .Ξ was viable, new21 was not" β PMID:21270388 |
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Download this section (compressed HTML)Q: Does cmc4 physically associate with an S. pombe SCO1/Cox-assembly ortholog, as its human counterpart does, or with a distinct IMS partner?
Q: Is cmc4 required for respiratory growth or cytochrome c oxidase activity/assembly in fission yeast, or is it fully dispensable as the viability of tam2Ξ and the lack of a respiratory phenotype in yeast CMC4 knockdown suggest?
Q: Does the meiotic (tam2) regulation of cmc4 reflect a meiosis- or sporulation-specific function, or merely coordinate expression of a mitochondrial protein with increased respiratory demand during differentiation?
Q: Does cmc4 bind a metal ion (e.g. copper) or participate directly in the MIA40-Erv1 disulfide relay as a substrate/accessory factor?
Experiment: Proximity labeling (BioID/TurboID) or affinity purification of endogenously tagged cmc4 in S. pombe under fermentative and respiratory (glycerol/ethanol) growth to identify direct IMS partners and test for association with Sco1/COX assembly factors.
Hypothesis: cmc4 associates with a COX II copper-delivery assembly factor (Sco1 ortholog) in the IMS, conserving the human CMC4-SCO1 interaction.
Experiment: Phenotypic characterization of a cmc4Ξ strain: growth on non-fermentable carbon sources, spectrophotometric cytochrome c oxidase activity and BN-PAGE COX assembly, and meiotic assays (sporulation efficiency, spore viability, meiotic progression timing).
Hypothesis: cmc4 loss produces a subtle, condition-specific respiratory and/or meiotic phenotype consistent with a non-essential accessory role in COX assembly.
Experiment: Biochemical characterization of recombinant cmc4: metal-binding assays (particularly Cu(I)) and thiol-disulfide/redox assays to test whether cmc4 is a metallochaperone or a MIA40-Erv1 relay accessory factor.
Hypothesis: cmc4 either binds a transition-metal ion or engages the disulfide relay beyond its own oxidative folding, defining its molecular activity.
What is not known β curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: cmc4 is a functionally dark gene: its molecular activity, its direct binding partner(s)/client in S. pombe, and its in vivo biological role are all undetermined. It is unresolved whether cmc4 acts in cytochrome c oxidase (COX II) assembly (as inferred from the human CMC4-SCO1 interaction) or has a distinct role connected to its meiotic (tam2) expression, and no cmc4Ξ phenotype has been reported.
OPEN BIOLOGY MF_DARK
What is known: What is firmly established: cmc4 (SPAC4F10.22 / tam2) is a small 70-residue mitochondrial intermembrane space protein of the twin CX9C / CHCH (CMC4) family, with two CX9C motifs forming a disulfide-bonded helical hairpin, imported and oxidatively folded by the MIA40-Erv1 disulfide relay. It is the sole fission-yeast member of a family conserved to human (CMC4, P56277). The human ortholog physically associates with the COX assembly factor SCO1, implicating the family in a non-essential branch of COX II sub-assembly, and the transcript is meiotically regulated in S. pombe (tam2), whose deletion is viable.
Significance: CMC4 is a conserved member of the twin CX9C protein family that collectively governs cytochrome c oxidase biogenesis and mitochondrial redox homeostasis, yet it is one of the least characterized members. Resolving its activity and in vivo role would close a genuine unknome gap and test whether the human COX II sub-assembly association is conserved and functionally important in a genetically tractable model.
What would resolve it: Affinity/proximity proteomics of tagged cmc4 in S. pombe to identify direct partners (test association with Sco1/Cox-assembly orthologs); recombinant metal-binding and thiol-disulfide assays to distinguish metallochaperone from disulfide-relay-accessory models; and phenotyping of cmc4Ξ for respiratory (COX activity on non-fermentable carbon) and meiotic (sporulation, spore viability, timing) defects.
Provenance (the field's own admissions):
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