GLT-1 (NCU01633) is a 532-residue, twelve-transmembrane-helix sugar porter of the major facilitator superfamily in Neurospora crassa. It is the low-affinity (Km about 18 mM), high-capacity glucose transporter of the dual-affinity glucose uptake system (historical system I), expressed under glucose-replete conditions, whereas the high-affinity HGT-1 and HGT-2 transporters supply uptake when glucose is limiting and can compensate for loss of GLT-1. Heterologous expression in a hexose-transporter-null yeast restores glucose uptake, and GFP-tagged GLT-1 was imaged in yeast and Neurospora, consistent with action at the plasma membrane. GLT-1 belongs to the PANTHER subfamily that includes the Aspergillus nidulans low-affinity glucose transporter MstE. Both lack the acidic TM1 residue that serves as the proton-binding site in proton-coupled sugar porters (Val48 in GLT-1), and whole-cell physiology describes system I as glucose diffusion, so GLT-1 is probably a facilitator rather than a sugar:proton symporter, although its energetics have not been measured directly.
Summary: GLT-1 transports glucose, but the inherited proton-coupling mechanism is over-specific.
Reason: PMID:28115989 assigns GLT-1 to historical low-affinity system I and describes system I as a glucose diffusion system. Primary structural work on the sugar-porter fold shows that XylE and GLUT1 are close homologs with different energetics and places XylE Asp27 versus GLUT1 Asn29 on the symporter/uniporter divide. The focused report verified that the current PTN000627328 symporter IBD is ancestral to the exact target leaf, so the IBA is procedurally current, but family-level sugar-porter ancestry does not establish conserved proton coupling. GLT-1 has non-protonatable Val48 at the TM1 position corresponding to XylE Asp27, while the high-affinity Neurospora HGT-1/HGT-2 paralogs retain acidic residues there. These sequence and physiological contrasts make GO:0005351 an over-specific mechanistic assertion for GLT-1; the core function should remain the mechanism-agnostic D-glucose transporter activity. An independent re-check (2026-10-21, glt-1-bioinformatics/RESULTS-rereview.md) reproduced Val48 and found that A. nidulans MstE, the characterised low-affinity glucose transporter in the same PANTHER subfamily (SF92), also has a non-acidic residue (Ser67) at this site, whereas HGT-1 Asp52 and HGT-2 Asp35 retain it. The PTN000627328 symporter IBD donors are SF6 Aspergillus transporters and yeast/fission-yeast members; MstE contributed to the carbohydrate-transport and membrane IBDs only, so the symporter inference reaches SF92 by descent, not from SF92 evidence. The single-residue diagnostic is asymmetric, so this is MARK_AS_OVER_ANNOTATED rather than REMOVE.
Propagation Review
Root cause:PROPAGATION BAD
Failure modes:FUNCTIONAL DIVERGENCEPSEUDO OR SUBACTIVITY LOSS
Sources checked:
PANTHER:PTN000627328 · PTN000627328 SUPPORTS SOURCE BUT NOT TARGET
Actual ancestry is verified, but the protein-specific Val48 loss of the symporter protonation-site residue and the HGT-1/HGT-2 acidic-residue contrast support loss of proton-coupled symport from GLT-1.
studies of XylE and GLUT1 suggest that the variation between an acidic residue (Asp27 in XylE) and a neutral one (Asn29 in GLUT1) is a key element for their mechanistic divergence
Structural comparison of the uniporter GLUT1 with its bacterial homologue XylE, a proton-coupled xylose symporter, allows examination of the transport mechanisms of both passive facilitators and active transporters.
In carbon-starved mycelia or mycelia incubated with galactose, a second galactose transport activity appeared which required energy, had a high affinity for galactose (Km = 0.7 mM), and was shown to be the same as glucose transport system II.
GLT-1 has a non-protonatable Val48 at this site, and the characterised low-affinity glucose transporter MstE from the same PANTHER subfamily (SF92) has a non-acidic Ser
IDA PMID:24581151 Transcriptional comparison of the filamentous fungus Neurosp...
NEW
Summary: Add the mechanism-agnostic D-glucose transporter activity directly established for GLT-1.
Reason: Heterologous expression and uptake assays establish GLT-1/NCU01633 as a D-glucose transporter, and this molecular-function term captures the demonstrated substrate specificity without importing the unsupported proton-coupling mechanism from the inherited GO:0005351 IBA.
Summary: GLT-1 transports carbohydrate; the substrate is experimentally resolved as D-glucose.
Reason: Heterologous complementation of a hexose-transporter-null yeast and radiolabeled glucose uptake (PMID:24581151, PMID:28115989) show GLT-1 transports D-glucose. The IBA arises at PTN000627328, an ancestor of the GLT-1 leaf, and the donor set includes A. nidulans MstE (Q5B0S0), a low-affinity glucose transporter in the same PANTHER subfamily (SF92). The inherited term is correct but less specific than the target evidence, so it is refined to D-glucose transmembrane transport, which GLT-1 already carries as a core process.
Propagation Review
Root cause:NO FAILURE CORE
Sources checked:
PANTHER:PTN000627328 · PTN000627328 SUPPORTS TRANSFER
Target leaf PTN002362095 descends from this IBD node via the SF92 subfamily node PTN002362094; the broad inference is true for GLT-1, and the SF92 member MstE is among the donors.
we assigned GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 and 98.97 ± 22.02 µM) to the low- and high-affinity glucose transport systems, respectively
Summary: GLT-1 is an integral membrane transporter; the supported location is the plasma membrane.
Reason: GLT-1 is a twelve-helix MFS sugar porter that mediates uptake of extracellular glucose, and GFP-tagged GLT-1 was imaged in yeast and in Neurospora (PMID:28115989). The SF92 subfamily relative MstE localises to the cell periphery consistent with the plasma membrane (PMID:16418173), and the membrane IBD at PTN000627328 is ancestral to the GLT-1 leaf. Membrane is correct but under-specified; the supported location is plasma membrane.
Propagation Review
Root cause:NO FAILURE CORE
Sources checked:
PANTHER:PTN000627328 · PTN000627328 SUPPORTS TRANSFER
Target leaf PTN002362095 descends from this IBD node via the SF92 subfamily node PTN002362094; membrane localisation is true for GLT-1 and is refined here to plasma membrane.
Summary: GLT-1 is an integral membrane transporter; the supported location is the plasma membrane.
Reason: This electronic membrane row (InterPro sugar-transporter signatures and UniProt subcellular-location mapping) is consistent with the IBA membrane row. The SF92 subfamily relative MstE localises to the cell periphery consistent with the plasma membrane (PMID:16418173), and the membrane IBD at PTN000627328 is ancestral to the GLT-1 leaf. Membrane is correct but under-specified; the supported location is plasma membrane.
Summary: GLT-1 is a transmembrane transporter whose substrate is experimentally resolved as D-glucose.
Reason: The InterPro sugar-transporter signatures correctly predict transmembrane transporter activity, and GLT-1-specific uptake assays resolve the substrate as D-glucose (PMID:24581151, PMID:28115989). The generic term is therefore refined to D-glucose transmembrane transporter activity, the mechanism-agnostic core activity proposed in this review.
we assigned GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 and 98.97 ± 22.02 µM) to the low- and high-affinity glucose transport systems, respectively
Summary: GLT-1 mediates carbohydrate transmembrane transport, specifically of D-glucose.
Reason: This row is inferred logically from the GO:0005351 IBA. Although the proton-coupling part of that term is judged over-annotated, the carbohydrate-transport part is independently supported by GLT-1 glucose uptake assays (PMID:24581151, PMID:28115989), so the process is true. It is refined to D-glucose transmembrane transport, the substrate-specific process.
we assigned GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 and 98.97 ± 22.02 µM) to the low- and high-affinity glucose transport systems, respectively
Summary: GLT-1 mediates transmembrane transport, specifically of D-glucose.
Reason: The InterPro-based transmembrane-transport inference is correct; GLT-1-specific uptake assays resolve the substrate as D-glucose (PMID:24581151, PMID:28115989), so the term is refined to D-glucose transmembrane transport.
we assigned GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 and 98.97 ± 22.02 µM) to the low- and high-affinity glucose transport systems, respectively
Summary: The proton-transport process row depends on an unsupported symport assumption.
Reason: GLT-1 has Val48 at the TM1 position corresponding to the XylE Asp27 protonation site, while the high-affinity Neurospora HGT-1/HGT-2 paralogs retain acidic residues there. This BP is electronically inferred only from GO:0005351 carbohydrate:proton symporter activity, so it inherits the same unsupported proton-coupling assumption. GLT-1-specific glucose uptake is established, but direct energetics have not been measured; once the GO:0005351 PAINT propagation is generalized to a mechanism-agnostic glucose transporter MF, this logical proton-transport row should disappear as well. The SF92 relative MstE likewise lacks the acidic TM1 residue (RESULTS-rereview.md).
studies of XylE and GLUT1 suggest that the variation between an acidic residue (Asp27 in XylE) and a neutral one (Asn29 in GLUT1) is a key element for their mechanistic divergence
we assigned GLT-1 (Km = 18.42 ± 3.38 mM) and HGT-1/-2 (Km = 16.13 ± 0.95 and 98.97 ± 22.02 µM) to the low- and high-affinity glucose transport systems, respectively
Q: Is there any GLT-1-specific proton-flux, voltage-clamp, or reconstitution datum not captured in the published Neurospora glucose-transporter literature, and if not should GLT-1's PAINT molecular-function row be generalized from carbohydrate:proton symporter activity to D-glucose transmembrane transporter activity?
Suggested Experiments
Experiment: Reconstitute purified GLT-1 into proteoliposomes and measure D-glucose uptake with and without imposed delta-pH and membrane-potential gradients; as an orthogonal test, express GLT-1 in Xenopus oocytes and assay whether glucose elicits a proton-dependent inward current.
Hypothesis: GLT-1 mediates low-affinity facilitated diffusion of D-glucose rather than proton-coupled glucose symport.
Type: Proteoliposome transport assay or oocyte electrophysiology
Knowledge Gaps
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: GLT-1 glucose uptake energetics are unresolved; no GLT-1-specific proton-flux, voltage-clamp, or reconstituted proteoliposome assay has directly distinguished facilitated diffusion from proton-coupled symport.
OPENBIOLOGYMF_DARK
What is known: GLT-1-specific radiolabeled uptake establishes low-affinity glucose transport, the TM1 Val48 scan argues against proton symport, and whole-cell Neurospora physiology historically distinguished low-affinity system I from high-affinity, energy-requiring system II without measuring GLT-1 alone.
Significance: Direct energetics would test the only remaining experimental caveat for replacing the inherited carbohydrate:proton symporter activity with a mechanism-agnostic D-glucose transporter activity.
What would resolve it: Reconstitute purified GLT-1 in proteoliposomes and measure glucose uptake with and without imposed pH and membrane-potential gradients, or express GLT-1 in Xenopus oocytes and test whether glucose evokes a proton-dependent inward current.
Provenance (the field's own admissions):
"1. **No direct GLT-1 energetics assay.** Checked: no EXP GO, no electrophysiology in the retrieved literature." — file:NEUCR/glt-1/glt-1-hypotheses/glucose-transport-and-proton-coupling/openscientist.md
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.
Review rationale: GLT-1 is the low-affinity D-glucose transporter of the N. crassa dual-affinity glucose uptake system; heterologous complementation of a hexose-transporter-null yeast and radiolabeled glucose uptake establish D-glucose transport (PMID:24581151, PMID:28115989). Transmembrane transporter activity is correct but less precise than the supported D-glucose transmembrane transporter activity and than the carbohydrate:proton symporter IBA the gene already carries in GOA, whose carbohydrate transport component is experimentally supported. The exact term is also already in GOA (InterPro IEA), so the prediction is not novel. This judgment concerns solute transport, not proton coupling.
Supporting Evidence:
PMID:24581151: "We conclude that NCU01633 is a D-glucose transporter of N. crassa, which was therefore named as D-glucose transporter 1 (GLT-1)."
PMID:28115989: "One low-affinity glucose transporter, GLT-1 (NCU01633), and two high-affinity transporters, HGT-1/-2 (NCU10021 and NCU04963), were identified as the major components of systems I and II, respectively."
Review rationale: GLT-1 is a twelve-helix MFS sugar porter that takes up extracellular glucose, and eGFP-tagged GLT-1 was imaged in both S. cerevisiae and N. crassa (PMID:28115989), supporting membrane residence; the supported location is the plasma membrane, inferred from the uptake function and the cell-periphery localisation of the same-subfamily relative MstE. Membrane is correct but less precise than that plasma-membrane assignment, and the exact term is already in GOA (IBA and IEA rows), so the prediction is not novel.
Supporting Evidence:
PMID:28115989: "Subcellular localization of HGT-1, HGT-2, and GLT-1 in Neurospora crassa."
PMID:24581151: "We conclude that NCU01633 is a D-glucose transporter of N. crassa, which was therefore named as D-glucose transporter 1 (GLT-1)."