Id3 (inhibitor of DNA binding 3; L-homeolog id3.L, with S-homeolog id3-b) is a small nuclear helix-loop-helix (HLH) protein that lacks the basic DNA-binding region of bHLH transcription factors. It heterodimerises with bHLH proteins such as E proteins (Tcf3/E12-E47) and acts as a dominant-negative inhibitor: it prevents myogenic and neurogenic bHLH/E-protein complexes from binding E-box DNA and so blocks their transcriptional activation (for example of MyoD- and NeuroD-driven programmes). It also binds and antagonises the bHLH-Orange repressor Hairy2/Hes4 and disrupts Stat3-FGFR4 complex formation. Id3 is expressed after the midblastula transition in pluripotent blastula ectoderm, then at the neural plate border and in premigratory and early migratory neural crest, and in eye, otic, myotome and other proliferating progenitor domains. It is induced by BMP, Notch and Myc. In the neural crest it is required to keep progenitors cycling, surviving and undifferentiated, and so is needed for formation of the multipotent neural crest progenitor pool; overexpression expands the crest domain and delays differentiation of crest derivatives. It therefore acts as a progenitor maintenance and competence factor downstream of the border signals, rather than as a border specifier or a DNA-binding crest fate specifier. The HLH inhibitor activity is ancestral to chordates (amphioxus Id is expressed in mesoderm and endoderm), whereas Id expression at the neural plate border and dorsal neural tube appeared early in the vertebrate lineage, which makes Id3 an example of co-option of a pre-existing regulator into the neural crest network.
Definition: The process by which neural plate border and premigratory neural crest progenitor cells are kept in an undifferentiated, proliferative and multipotent state until neural crest specification.
Justification: GO:0014029 neural crest formation, which these factors currently carry, is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md). Competence factors (Myc, Id3, Hairy2, Pou5f3/Oct25) are required for crest to form, but loss causes progenitor arrest or death rather than a change of fate, and sustained activity blocks differentiation, so neither crest cell differentiation nor fate specification describes their role. GO:0019827 stem cell population maintenance is the parent; the new term says which progenitor population is maintained.
Parent term: stem cell population maintenance
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0000122 negative regulation of transcription by RNA polymerase II | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference that Id-family proteins negatively regulate RNA polymerase II transcription by sequestering bHLH activators. Q91399 appears among the descendant evidences because it has its own experimental annotation for this term. Reason: Id proteins act as dominant-negative HLH inhibitors of bHLH transcriptional activators across vertebrates, and Xenopus Id3 itself blocks MyoD-driven E-box transactivation in embryonic tissue. The node placement is sound and the term is at an appropriate level for the process outcome of Id3 activity. Supporting Evidence: PMID:7734394 disrupts binding of myogenic factor/E-protein complexes to DNA in vitro and inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0000122 negative regulation of transcription by RNA polymerase II | IDA PMID:14651922 Cloning and characterization of Xenopus Id4 reveals differin... | ACCEPT | Summary: Liu and Harland compared Xenopus Id2, Id3 and Id4 in animal caps and in vivo and found that Id3 selectively blocks the activity of the neurogenic bHLH factor NeuroD. Reason: Inhibition of NeuroD-driven activity is inhibition of a bHLH transcriptional activator, which is the canonical Id mechanism leading to reduced Pol II transcription of bHLH target genes. Abstract-only in cache; the curator read the full text and the result is consistent with the family mechanism. Supporting Evidence: PMID:14651922 Id4 is able to inhibit the activity all these bHLH molecules, Id2 inhibits MyoD and neuroD, while Id3 blocks only neuroD activity in our assays. |
| GO:0000122 negative regulation of transcription by RNA polymerase II | IDA PMID:7619724 Id gene activity during Xenopus embryogenesis. | ACCEPT | Summary: Zhang et al. cloned XIdI (Id3) and described its expression; UniProt cites this paper for function. The abstract frames Ids as inhibitors of bHLH activity. Reason: The cached abstract is expression-focused and does not itself describe the assay the curator used, but the asserted process is correct for Id3 and is independently supported (PMID:7734394). Deferring to the curator who read the full text. Supporting Evidence: PMID:7619724 The activity of bHLH transcription factors that are involved in cell determination and differentiation is inhibited by Ids, HLH proteins lacking the basic amino acid sequence element. PMID:7734394 inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0000122 negative regulation of transcription by RNA polymerase II | IDA PMID:7734394 XIdx, a dominant negative regulator of bHLH function in earl... | ACCEPT | Summary: Wilson and Mohun showed that XIdx (Id3) disrupts myogenic factor/E-protein DNA binding and inhibits MyoD transactivation of the E-box regulated cardiac actin gene in embryonic tissue. Reason: Direct functional evidence that Xenopus Id3 represses bHLH-dependent Pol II transcription by a dominant-negative mechanism. This is the core process outcome of Id3 molecular activity. Supporting Evidence: PMID:7734394 XIdx, which disrupts binding of myogenic factor/E-protein complexes to DNA in vitro and inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue. |
| GO:0000122 negative regulation of transcription by RNA polymerase II | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic annotation from the DNA-binding protein inhibitor family (IPR026052) and PANTHER node; same term as the IBA and IDA rows. Reason: Consistent with the experimental and phylogenetic evidence for Id3 as a dominant-negative inhibitor of bHLH transcriptional activators. Supporting Evidence: PMID:7734394 inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0000122 negative regulation of transcription by RNA polymerase II | IPI PMID:18721802 Hairy2-Id3 interactions play an essential role in Xenopus ne... | ACCEPT | Summary: Based on the physical interaction of Id3 with Hairy2 (Hes4-b). In this paper Id3 binds Hairy2 and negatively regulates Hairy2 activity. Reason: The gene-term pair is correct: Id3 negatively regulates Pol II transcription (PMID:7734394). This particular row is weakly grounded, though. Hairy2/Hes4 is itself a transcriptional repressor, so Id3 antagonism of Hairy2 would be expected to relieve repression at Hairy2 targets. A binding (IPI) experiment does not by itself establish the direction of a transcriptional outcome either. The full text is not cached, so the readout the curator used cannot be checked, and the row is accepted on the strength of the gene-level evidence. Supporting Evidence: PMID:18721802 Knockdown and rescue experiments indicate that Id3 protein, which physically interacts with Hairy2, negatively regulates Hairy2 activity. PMID:7734394 inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0003714 transcription corepressor activity | IBA GO_REF:0000033 | MODIFY | Summary: Phylogenetic inference of corepressor activity for the Id family (seeded from Drosophila extramacrochaetae). Reason: The family membership is right but the molecular function term is not. GO defines a corepressor as acting by binding a DNA-binding transcription factor at a specific genomic locus. Id proteins work the other way round: they lack a basic region, heterodimerise with bHLH proteins off DNA, and prevent those complexes from binding E-boxes. Xenopus Id3 disrupts myogenic factor/E-protein DNA binding and also disrupts Stat3-FGFR4 complex formation. GO:0140416 transcription regulator inhibitor activity describes this sequestration mechanism and is the term used for experimentally characterised mammalian ID1, ID2 and ID4. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000919548 SUPPORTS TRANSFER Id-family ancestral node. The inhibitory regulator role is correctly inherited by Id3, but the corepressor term implies action as a cofactor on DNA-bound TFs; the inherited activity is better typed as transcription regulator inhibitor activity. FB:FBgn0000575 SUPPORTS TRANSFER Drosophila emc, an HLH protein without a basic region that antagonises proneural bHLH factors; supports the inhibitory function, not a DNA-bound corepressor mechanism. Proposed replacements: transcription regulator inhibitor activity Supporting Evidence: PMID:7734394 disrupts binding of myogenic factor/E-protein complexes to DNA in vitro PMID:7619724 The activity of bHLH transcription factors that are involved in cell determination and differentiation is inhibited by Ids, HLH proteins lacking the basic amino acid sequence element. file:XENLA/id3-a/id3-a-deep-research-falcon.md Id proteins retain an HLH dimerization region but lack the basic DNA-binding region characteristic of basic HLH (bHLH) transcription factors. |
| GO:0005515 protein binding | IPI PMID:19851287 Self-regulation of Stat3 activity coordinates cell-cycle pro... | REMOVE | Summary: Interaction of Id3 with Stat3 (stat3.1, Q9PVX8). Id3 disrupts Stat3-FGFR4 complex formation and so limits FGF-driven Stat3 activation in the ectoderm. Reason: Generic protein binding is uninformative. The interaction is real and functionally meaningful: it is an instance of Id3 inhibiting a transcription regulator by sequestration. That content is captured by GO:0140416 transcription regulator inhibitor activity in core_functions and the MODIFY above. Removal does not mean the interaction is false. Supporting Evidence: PMID:19851287 Stat3 activation is also modulated by Hairy2 and Id3 proteins that, respectively, facilitate and disrupt Stat3-FGFR4 complex formation. |
| GO:0005634 nucleus | IBA GO_REF:0000033 | ACCEPT | Summary: Phylogenetic inference that Id proteins act in the nucleus. Reason: Id3 inhibits DNA binding and transactivation by nuclear bHLH factors, so the nucleus is its site of action. No Xenopus protein-localisation data exist, but the IBA is well grounded in the family. Supporting Evidence: PMID:7734394 disrupts binding of myogenic factor/E-protein complexes to DNA in vitro and inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0005634 nucleus | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic nuclear localisation from UniProt subcellular location (by similarity to human ID3) and the PANTHER node. Reason: Consistent with the IBA and with the nuclear mechanism of Id3 action. |
| GO:0005737 cytoplasm | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro2GO cytoplasm annotation for the DNA-binding protein inhibitor family. Reason: Some Id proteins (for example ID1 and ID2) are seen in the cytoplasm and small HLH proteins shuttle, so cytoplasmic presence is plausible. However, there is no Xenopus Id3 protein localisation data, and the Id3 activities that matter (inhibiting bHLH DNA binding, antagonising Hairy2) are nuclear. Id3 disruption of Stat3-FGFR4 complexes could take place in the cytoplasm, but this was not localised. Kept as non-core. Supporting Evidence: PMID:19851287 Stat3 activation is also modulated by Hairy2 and Id3 proteins that, respectively, facilitate and disrupt Stat3-FGFR4 complex formation. |
| GO:0014029 neural crest formation | IMP PMID:15769946 To proliferate or to die: role of Id3 in cell cycle progress... | MODIFY | Summary: Kee and Bronner-Fraser: Id3 is expressed at the neural plate border; morpholino depletion removes neural crest precursors through cell-cycle arrest and death of the progenitor pool, not through a fate switch; overexpression expands the crest. Reason: Loss of function, gain of function and expression all point to Id3 being needed to build the neural crest progenitor pool. The authors state that this is independent of cell fate determination. Fate specification (GO:0014036) or fate commitment (GO:0014034) would overstate the role. Id3 acts as a progenitor maintenance and competence factor, downstream of the border signals. GO:0014029 is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md): its definition describes forming the neural plate border region while its placement is under epithelial to mesenchymal transition. This gene's role is keeping crest progenitors undifferentiated and competent, captured by the proposed term 'neural crest progenitor maintenance' (NTR). Proposed replacements: neural crest progenitor maintenance Supporting Evidence: PMID:15769946 Morpholino oligonucleotide-mediated depletion of Id3 results in the absence of neural crest precursors and a resultant loss of neural crest derivatives. PMID:15769946 This appears to be mediated by cell cycle inhibition followed by cell death of the neural crest progenitor pool, rather than a cell fate switch. PMID:15769946 overexpression of Id3 increases cell proliferation and results in expansion of the neural crest domain |
| GO:0014029 neural crest formation | IMP PMID:15772131 Xenopus Id3 is required downstream of Myc for the formation ... | MODIFY | Summary: Light et al.: Id3 is a Myc target; knockdown gives embryos without neural crest, and forced expression maintains progenitor markers and blocks differentiation of crest derivatives. Reason: Independent loss-of-function evidence from a second lab, with gain-of-function data showing that Id3 keeps cells in the multipotent progenitor state. This supports maintaining the progenitor pool rather than fate specification. GO:0014029 is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md): its definition describes forming the neural plate border region while its placement is under epithelial to mesenchymal transition. This gene's role is keeping crest progenitors undifferentiated and competent, captured by the proposed term 'neural crest progenitor maintenance' (NTR). Proposed replacements: neural crest progenitor maintenance Supporting Evidence: PMID:15772131 the small HLH protein Id3 is a Myc target that plays an essential role in the formation and maintenance of neural crest stem cells PMID:15772131 A morpholino-mediated 'knockdown' of Id3 protein results in embryos that lack neural crest. |
| GO:0014029 neural crest formation | IMP PMID:18710660 Hairy2 functions through both DNA-binding and non DNA-bindin... | MODIFY | Summary: Nichane et al. (Hairy2 DNA-binding and non-DNA-binding mechanisms). The abstract concerns Hairy2, Delta1 and Stat3; UniProt cites this paper for Id3 function and induction. Reason: The cached abstract does not mention Id3, but the full text (not cached) assays Id3 in the Hairy2/Stat3/Delta1 border circuit, and the companion paper (PMID:18721802) shows Id3 is an essential neural crest regulator. The term is correct for Id3, so the curator's IMP is accepted. GO:0014029 is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md): its definition describes forming the neural plate border region while its placement is under epithelial to mesenchymal transition. This gene's role is keeping crest progenitors undifferentiated and competent, captured by the proposed term 'neural crest progenitor maintenance' (NTR). Proposed replacements: neural crest progenitor maintenance Supporting Evidence: PMID:18721802 We show that Hairy2 downregulates Id3, another essential HLH neural crest regulator, through attenuation of BMP signaling. |
| GO:0014029 neural crest formation | IMP PMID:18721802 Hairy2-Id3 interactions play an essential role in Xenopus ne... | MODIFY | Summary: Nichane et al.: Id3 is an essential HLH neural crest regulator. It binds and antagonises Hairy2 but is required for Hairy2 to promote neural crest formation. Reason: Knockdown and rescue data from a third lab support the requirement of Id3 for neural crest progenitor formation. The paper uses the word specification, but the combined evidence (cell-cycle and survival phenotype, no fate switch) supports progenitor maintenance rather than GO:0014036 fate specification. GO:0014029 is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md): its definition describes forming the neural plate border region while its placement is under epithelial to mesenchymal transition. This gene's role is keeping crest progenitors undifferentiated and competent, captured by the proposed term 'neural crest progenitor maintenance' (NTR). Proposed replacements: neural crest progenitor maintenance Supporting Evidence: PMID:18721802 However, Id3 is required to allow Hairy2 to promote neural crest formation. |
| GO:0014029 neural crest formation | IMP PMID:19851287 Self-regulation of Stat3 activity coordinates cell-cycle pro... | MODIFY | Summary: Nichane et al. 2010: Stat3 activity downstream of FGF coordinates cell-cycle progression and neural crest specification under positive (Hairy2) and negative (Id3) feedback. Reason: Places Id3 in the FGF-Stat3 feedback loop that balances proliferation and differentiation in neural crest progenitors, in line with the other three perturbation studies. GO:0014029 is proposed for obsoletion (projects/NEURAL_CREST_FORMATION_OBSOLETION.md): its definition describes forming the neural plate border region while its placement is under epithelial to mesenchymal transition. This gene's role is keeping crest progenitors undifferentiated and competent, captured by the proposed term 'neural crest progenitor maintenance' (NTR). Proposed replacements: neural crest progenitor maintenance Supporting Evidence: PMID:19851287 Stat3, downstream of FGFs and under the positive and negative feedback regulation of Hairy2 and Id3, plays an essential role in the coordination of cell-cycle progression and differentiation during NC specification |
| GO:0030154 cell differentiation | IEA GO_REF:0000118 | MODIFY | Summary: TreeGrafter annotation to the very general term cell differentiation. Reason: Id proteins are inhibitors of differentiation. In Xenopus, forced Id3 expression maintains progenitor markers and blocks differentiation of neural crest derivatives, and Id3 blocks NeuroD activity. The direction is known, so the annotation should be to negative regulation of cell differentiation rather than the neutral parent. Proposed replacements: negative regulation of cell differentiation Supporting Evidence: PMID:15772131 forced expression of Id3 maintains the expression of markers of the neural crest progenitor state beyond the time when they would normally be downregulated and blocks the differentiation of neural crest derivatives |
| GO:0030182 neuron differentiation | IBA GO_REF:0000033 | MODIFY | Summary: Phylogenetic inference that Id proteins take part in neuron differentiation. Reason: Id-family involvement in neurogenesis is real, but its direction is inhibitory: Id proteins hold neural progenitors back from proneural bHLH-driven differentiation. For Xenopus Id3 the direct evidence is that it blocks NeuroD activity. Negative regulation of neuron differentiation captures this better. It remains non-core for Id3, whose best-characterised role is in neural crest progenitors. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000919548 SUPPORTS TRANSFER Id-family ancestral node. The neurogenesis role is inherited, but Id proteins act as negative regulators of proneural bHLH-driven differentiation; the neutral process term loses the sign that the family evidence supports. Proposed replacements: negative regulation of neuron differentiation Supporting Evidence: PMID:14651922 Id proteins can differentially inhibit the activities of neurogenin and neuroD, both neurogenic bHLH molecules and MyoD, a myogenic bHLH protein. PMID:14651922 Id3 blocks only neuroD activity in our assays |
| GO:0032922 circadian regulation of gene expression | IEA GO_REF:0000118 | REMOVE | Summary: TreeGrafter propagation of a circadian role to Xenopus Id3. Reason: Experimental circadian evidence in the Id family is for Id2 (mouse Id2 IMP; human ID2 by ISS). Human ID3 itself has no circadian annotation, and there is no Xenopus Id3 evidence for a clock role. The UniProt function text states only that it "may play a role" by similarity. This is an over-propagation from an Id2-specific function to an Id3 subfamily member. Supporting Evidence: file:XENLA/id3-a/id3-a-deep-research-falcon.md The most defensible functional annotation is: **an HLH-mediated antagonist of bHLH transcriptional activity that, in the Xenopus Id3 experimental literature, sustains proliferation and survival of neural-crest progenitors and integrates developmentally restricted BMP/Notch-associated inputs.** |
| GO:0043392 negative regulation of DNA binding | IDA PMID:7619724 Id gene activity during Xenopus embryogenesis. | ACCEPT | Summary: Zhang et al. (XIdI cloning). The abstract describes Ids as HLH proteins lacking the basic region that inhibit bHLH activity; the DNA-binding assay itself is not described in the cached abstract. Reason: Inhibition of bHLH DNA binding is the defining mechanism of Id proteins and is shown directly for Xenopus Id3 in PMID:7734394. The full text is not cached, so the curator's IDA is accepted. Supporting Evidence: PMID:7619724 The activity of bHLH transcription factors that are involved in cell determination and differentiation is inhibited by Ids, HLH proteins lacking the basic amino acid sequence element. |
| GO:0043392 negative regulation of DNA binding | IDA PMID:7734394 XIdx, a dominant negative regulator of bHLH function in earl... | ACCEPT | Summary: XIdx (Id3) disrupts binding of myogenic factor/E-protein complexes to DNA in vitro. Reason: Direct biochemical evidence of the core mechanism: Id3 reduces bHLH DNA binding by sequestering partners into non-DNA-binding heterodimers. The V59N HLH mutant, which reduces this inhibition, supports specificity. Supporting Evidence: PMID:7734394 disrupts binding of myogenic factor/E-protein complexes to DNA in vitro |
| GO:0043425 bHLH transcription factor binding | IPI PMID:18721802 Hairy2-Id3 interactions play an essential role in Xenopus ne... | ACCEPT | Summary: Id3 physically interacts with the bHLH-Orange repressor Hairy2 (Hes4-b, Q90VV1) and negatively regulates its activity. Reason: Hairy2 is a bHLH transcription factor and the interaction is functionally relevant at the neural plate border. bHLH transcription factor binding is an informative partner-class term for an Id protein. Supporting Evidence: PMID:18721802 Id3 protein, which physically interacts with Hairy2, negatively regulates Hairy2 activity |
| GO:0043425 bHLH transcription factor binding | IPI PMID:7619724 Id gene activity during Xenopus embryogenesis. | ACCEPT | Summary: Interaction with the E protein Tcf3 (XE12/XE47, Q01978), the canonical Id partner. Reason: Heterodimerisation with E proteins is the canonical Id mechanism. The full text is not cached, but the interaction with an E protein is fully consistent with the family biology and with PMID:7734394. Supporting Evidence: PMID:7619724 The activity of bHLH transcription factors that are involved in cell determination and differentiation is inhibited by Ids, HLH proteins lacking the basic amino acid sequence element. |
| GO:0043425 bHLH transcription factor binding | IPI PMID:7734394 XIdx, a dominant negative regulator of bHLH function in earl... | ACCEPT | Summary: XIdx (Id3) interacts with E protein Tcf3 (XE12/XE47) and disrupts myogenic factor/E-protein DNA binding. Reason: Binding of bHLH E proteins underlies the dominant-negative activity of Id3 and is a core part of its molecular function. Supporting Evidence: PMID:7734394 disrupts binding of myogenic factor/E-protein complexes to DNA in vitro |
| GO:0045892 negative regulation of DNA-templated transcription | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO annotation; parent of negative regulation of transcription by RNA polymerase II. Reason: Correct, although less specific than GO:0000122, which is also annotated. Supporting Evidence: PMID:7734394 inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0045892 negative regulation of DNA-templated transcription | ISS GO_REF:0000024 | ACCEPT | Summary: Transferred by sequence similarity from human ID3 (P41133 is listed as source). Reason: Consistent with the direct Xenopus evidence; correct but broader than GO:0000122. Supporting Evidence: PMID:7734394 inhibits transactivation of the E-box regulated cardiac actin gene by MyoD in embryonic tissue |
| GO:0046983 protein dimerization activity | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro2GO annotation from the HLH domain. Reason: Id3 works by HLH-mediated heterodimerisation with bHLH partners, and the V59N HLH mutation reduces its inhibitory activity. The term is generic, but it is correct for an HLH protein whose activity depends on dimerisation. Partner class and functional consequence are captured more informatively by bHLH transcription factor binding and transcription regulator inhibitor activity. Supporting Evidence: file:XENLA/id3-a/id3-a-deep-research-falcon.md Id proteins retain an HLH dimerization region but lack the basic DNA-binding region characteristic of basic HLH (bHLH) transcription factors. |
| GO:0051726 regulation of cell cycle | IMP PMID:15769946 To proliferate or to die: role of Id3 in cell cycle progress... | MODIFY | Summary: Id3 depletion causes cell-cycle inhibition (fewer mitotic cells, ectopic p27Xic1) in neural crest progenitors, and overexpression increases proliferation. Reason: The direction is established by loss of function (cell-cycle arrest) and gain of function (increased proliferation), so positive regulation of cell cycle is more precise. This proliferative role in neural crest progenitors is central to how Id3 supports neural crest formation, and matches the conserved role of Id proteins in keeping progenitors cycling. Proposed replacements: positive regulation of cell cycle Supporting Evidence: PMID:15769946 the helix-loop-helix transcriptional regulator Id3 has a novel role in cell cycle progression and survival of neural crest progenitors in Xenopus PMID:15769946 overexpression of Id3 increases cell proliferation and results in expansion of the neural crest domain file:XENLA/id3-a/id3-a-deep-research-falcon.md Conversely, Id3 overexpression increased BrdU labeling and expanded the neural-crest-marker domain. |
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Download this section (compressed HTML)Q: GO has no neural plate border formation/specification term. Id3 is expressed at the border but acts downstream of border signals, as a progenitor maintenance factor. If a border term is created, should Id3 be excluded from it (in contrast to Pax3, Zic1, Msx1, Tfap2a and Hairy2), so that the term keeps the border-specifier layer distinct?
Q: Is GO:0019827 stem cell population maintenance (or a neural crest-specific progenitor maintenance term) a better description of the Id3 role than neural crest formation, given that forced Id3 maintains progenitor markers and that Id3 is shared with the blastula and ES-cell potency programme? Is there Xenopus evidence that Id3 is required in blastula cells, rather than only co-expressed with Oct/Sox/Vent?
Q: Which bHLH partner(s) does Id3 sequester in neural crest progenitors to prevent p27Xic1 induction and cell-cycle exit? Are E proteins the relevant partners, or is Hairy2/Stat3 antagonism sufficient to explain the phenotype?
Q: Do id3.L (Q91399) and id3.S (Q7SZ28) differ in expression or activity? All experimental annotations are attached to id3-a, while id3-b carries ISS copies; the morpholinos used most likely targeted both homeologs.
Q: Amphioxus Id is expressed in mesoderm and endoderm but not at the neural plate border, while lamprey Id is expressed in neural crest. Was Id co-opted into the border by acquiring BMP/Notch/Myc-responsive enhancers in the vertebrate ancestor, and does amphioxus Id protein retain the same bHLH-inhibitory activity (that is, a purely cis-regulatory co-option)?
Experiment: In Id3 morphant or CRISPR F0 Xenopus embryos, rescue with wild-type Id3, the V59N HLH mutant, or Id3 variants engineered to bind E proteins but not Hairy2 (or the reverse). Score proliferation (pH3, EdU), p27Xic1 expression and neural crest markers, and use proximity labelling in neural plate border explants to identify the Id3 partners in vivo.
Hypothesis: Id3 maintains neural crest progenitors by sequestering E proteins and thereby preventing activation of cell-cycle inhibitors such as p27Xic1.
Experiment: Test whether amphioxus Id and lamprey Id protein rescue Xenopus Id3 loss of function in the neural crest, and compare Xenopus id3 neural plate border enhancers (BMP response element, Su(H) sites, Myc E-boxes) with amphioxus Id regulatory regions in reporter assays in Xenopus and chick.
Hypothesis: The Id3 neural crest function is cis-regulatory co-option of an ancestral HLH inhibitor.
Experiment: Generate homeolog-specific CRISPR mutants of id3.L and id3.S, singly and in combination, and assay neural crest marker expression and progenitor proliferation.
Hypothesis: id3.L and id3.S contribute redundantly to neural crest progenitor maintenance.
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