VAM10 (YOR068C) encodes a small (114 aa) peripheral vacuolar-membrane protein of Saccharomyces cerevisiae required for homotypic (non-autophagic) vacuole fusion. Its gene is nested within the VPS5 locus on the opposite DNA strand. Vam10p acts early in the vacuole fusion cascade at a priming step that is independent of the canonical Sec18p (NSF)/Sec17p (alpha-SNAP) SNARE-disassembly machinery, and this Vam10p-dependent priming is a prerequisite for the subsequent Ypt7p (Rab GTPase)- dependent tethering of vacuoles. Deletion causes vacuole fragmentation, and recombinant Vam10p stimulates fusion of purified vacuoles in vitro while anti-Vam10p antibody blocks it. The protein has no recognizable catalytic or binding domain and no strong orthology outside fungi, and its molecular activity and direct binding partners remain undefined.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005774 vacuolar membrane | IEA GO_REF:0000044 | ACCEPT | Summary: Electronic Subcellular Location mapping placing Vam10p at the vacuolar membrane. Consistent with the experimentally-informed IC call to the fungal-type vacuole membrane and with the protein's role at the vacuole surface during fusion. Reason: Correct location and supported by an independent, more specific IC annotation (GO:0000329) from the primary functional study. Kept as a valid but non-core supporting cellular-component annotation; the fungal-type vacuole membrane term is preferred for representing the site of action. |
| GO:0000329 fungal-type vacuole membrane | IC PMID:12748377 Vam10p defines a Sec18p-independent step of priming that all... | ACCEPT | Summary: Curator-inferred (IC) localization to the fungal-type vacuole membrane, based on the vacuole-fusion function reported by Kato & Wickner (2003). Species-appropriate and the informative cellular-component annotation for this protein. Reason: The functional study demonstrates Vam10p acts on purified vacuoles and restores tethering to vacuoles from a vam10-delta strain, and UniProt independently curates a vacuole-membrane (peripheral) location; the fungal-type vacuole membrane is the correct organelle-specific term for a Saccharomyces protein. Supports the core-function location. Supporting Evidence: PMID:12748377 Pure Vam10p restores normal, Ypt7p-dependent tethering to vacuoles from a vam10Delta strain. |
| GO:0003674 molecular_function | ND GO_REF:0000015 | ACCEPT | Summary: Root molecular_function annotation with the ND (No biological Data available) evidence code, marking that no specific molecular activity has been experimentally or computationally assigned to Vam10p. Reason: This is an honest placeholder that correctly reflects the genuine molecular-function gap for this protein: Vam10p has no recognizable catalytic or binding domain, no informative orthology, and no biochemically defined activity. It should NOT be replaced with a specific molecular-function term; the missing activity is recorded under knowledge_gaps instead. Not a core function. |
| GO:0042144 vacuole fusion, non-autophagic | IMP PMID:12748377 Vam10p defines a Sec18p-independent step of priming that all... | ACCEPT | Summary: Involvement in homotypic (non-autophagic) vacuole fusion, from the deletion phenotype (vacuole fragmentation) and in-vitro reconstitution (recombinant Vam10p stimulates fusion of purified vacuoles; anti-Vam10p antibody blocks it). This is the core, well-supported biological role of the gene. Reason: Directly supported by loss-of-function (in-vivo fragmentation; antibody inhibition of in-vitro fusion) and gain-of-function (recombinant-protein stimulation) evidence in the same study. Represents the core biological process of VAM10. Supporting Evidence: PMID:12748377 VAM10 deletion causes vacuole fragmentation in vivo. The in vitro fusion of purified yeast vacuoles is stimulated by recombinant Vam10p and blocked by antibody to Vam10p. |
Loading supporting content…
Download this section (compressed HTML)Q: What is the molecular activity of Vam10p during the Sec18p-independent priming step that licenses Ypt7p-dependent tethering?
Suggested experts: William Wickner
Q: Does Vam10p act directly on the Rab GTPase Ypt7p, on the HOPS complex, on vacuolar SNAREs, or on the vacuolar lipid bilayer?
Suggested experts: Michiko Kato, William Wickner
Experiment: Affinity-purify or crosslink epitope-tagged Vam10p from isolated vacuoles and identify co-purifying proteins by mass spectrometry; test candidate interactions (Ypt7p, HOPS subunits, vacuolar SNAREs) by directed pulldowns, and assay lipid binding of purified recombinant Vam10p.
Hypothesis: Vam10p binds a specific vacuolar component (Ypt7p, a HOPS subunit, a SNARE, or a lipid) to promote the pre-tethering priming step.
Type: interaction proteomics / biochemical binding assays
Experiment: Use the in-vitro homotypic vacuole fusion assay with order-of-addition and stage-specific inhibitors to place the Vam10p requirement precisely relative to Sec18p-mediated priming and Ypt7p/HOPS-mediated tethering, and characterize what Vam10p acts on during the reaction.
Hypothesis: The Vam10p-dependent priming reaction produces a defined change on the vacuole membrane required before Ypt7p-dependent tethering.
Type: in-vitro reconstituted membrane fusion
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: The mechanism of the Sec18p-independent priming step defined by Vam10p is unknown: what change in the vacuolar membrane or its protein/lipid composition Vam10p produces to license subsequent Ypt7p-dependent tethering has not been determined.
OPEN BIOLOGY MF_DARK
What is known: Canonical priming (Sec18p/NSF- and Sec17p/alpha-SNAP-driven SNARE-complex disassembly) is well characterized. Vam10p provides a functional marker for a separate, early, Sec18p-independent priming activity, but the reaction it catalyzes or promotes is not defined.
Significance: A second priming activity distinct from NSF/alpha-SNAP would refine the textbook priming-tethering-docking-fusion model of homotypic membrane fusion.
What would resolve it: In-vitro dissection of the Vam10p-dependent priming reaction (order-of-addition, substrate/product analysis) to define what Vam10p acts on and what it produces.
Provenance (the field's own admissions):
Gap: Whether Vam10p has orthologs with a conserved function beyond Saccharomyces (in other fungi or in higher eukaryotes) is uncharacterized, so it is unknown whether the Sec18p-independent priming role is a general feature of membrane fusion or a yeast-specific adaptation.
OPEN BIOLOGYCURATION BP_DARK
What is known: Vam10p is a small protein with no recognizable domain and only weak/singleton orthology signals in HOGENOM/InParanoid/OrthoDB; no functionally validated ortholog has been reported.
Significance: Establishing conservation would indicate whether the priming activity is a widely used mechanism worth mapping onto human membrane-fusion pathways.
What would resolve it: Phylogenetic profiling and functional complementation tests of candidate fungal homologs; structural comparison of the predicted fold against known fusion factors.
Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)Loading supporting content…
Download this section (compressed HTML)