Dll3 (Delta-like 3) is a divergent, vertebrate-specific member of the DSL (Delta/Serrate/LAG-2) family of Notch ligands, a type I transmembrane protein with an N-terminal MNNL domain and six EGF-like repeats but a degenerate DSL domain. Unlike Dll1 and Dll4, Dll3 does not bind Notch receptors in trans and does not activate Notch signalling. Instead it acts cell autonomously as a Notch antagonist (cis-inhibition): endogenous Dll3 resides predominantly in the Golgi apparatus and late endocytic compartments, where it associates with newly synthesised Notch1 and is proposed to divert it to lysosomal degradation, attenuating Notch signalling induced by other DSL ligands. Dll3 is expressed in the paraxial/presomitic mesoderm and in differentiating neurons; in mouse, loss of Dll3 (pudgy and targeted alleles) disrupts the segmentation clock and somite boundary and rostro-caudal polarity formation, producing severe vertebral and rib malformations, and DLL3 mutations cause human spondylocostal dysostosis type 1.
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0001501 skeletal system development | IEA GO_REF:0000107 | KEEP AS NON CORE | Summary: Dll3 null (neo) and pudgy alleles cause severe vertebral and rib (axial skeletal) defects. Reason: The skeletal phenotype is a downstream consequence of defective somite segmentation; Dll3 does not act directly in skeletal development. Supporting Evidence: PMID:11923214 the developmental origins of the skeletal defects lie in delayed and irregular somite formation, which results in the perturbation of anteroposterior somite polarity. |
| GO:0001501 skeletal system development | IMP PMID:11923214 Axial skeletal defects caused by mutation in the spondylocos... | KEEP AS NON CORE | Summary: Dll3 null (neo) and pudgy alleles cause severe vertebral and rib (axial skeletal) defects. Reason: The skeletal phenotype is a downstream consequence of defective somite segmentation; Dll3 does not act directly in skeletal development. Supporting Evidence: PMID:11923214 the developmental origins of the skeletal defects lie in delayed and irregular somite formation, which results in the perturbation of anteroposterior somite polarity. |
| GO:0001501 skeletal system development | ISO GO_REF:0000119 | KEEP AS NON CORE | Summary: Dll3 null (neo) and pudgy alleles cause severe vertebral and rib (axial skeletal) defects. Reason: The skeletal phenotype is a downstream consequence of defective somite segmentation; Dll3 does not act directly in skeletal development. Supporting Evidence: PMID:11923214 the developmental origins of the skeletal defects lie in delayed and irregular somite formation, which results in the perturbation of anteroposterior somite polarity. |
| GO:0001701 in utero embryonic development | NAS PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | MARK AS OVER ANNOTATED | Summary: NAS from the Dll3 expression-cloning paper, which described expression during gastrulation and early organogenesis. Reason: Expression during embryogenesis does not establish a role in in utero embryonic development in general; Dll3 mutant phenotypes are confined to somite segmentation, and pudgy/null mice survive to birth and beyond. The informative process is somitogenesis. Supporting Evidence: PMID:9272948 An extensive expression study during gastrulation and early organogenesis in the mouse reveals a diverse and dynamic pattern of expression. |
| GO:0001709 cell fate determination | NAS PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | MARK AS OVER ANNOTATED | Summary: NAS from the expression paper hypothesising roles in neuronal differentiation. Reason: Speculative, expression-based statement. Later work shows Dll3 attenuates Notch signalling cell-autonomously rather than determining fate as a signalling ligand; no loss-of-function evidence supports a cell fate determination role. Supporting Evidence: PMID:9272948 We hypothesise that Dll1 is involved in the release of cells from the precursor population and that Dll3 is required later to divert neurons along a specific differentiation pathway. |
| GO:0001756 somitogenesis | IDA PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | ACCEPT | Summary: Dll3 is expressed in the PSM and Dll3 loss (pudgy, targeted null) disrupts somite formation, segmentation-clock gene expression and somite polarity. Reason: Somitogenesis is the principal in vivo process requiring Dll3 (core process). Supporting Evidence: PMID:15170697 We report here that the Dll3(pu) mutation has different effects on the expression of cycling (Lfng and Hes7) and stage-specific genes (Hey3 and Mesp2). PMID:9662403 the pu mutation disrupts the proper formation of morphological borders in early somite formation and of rostral-caudal compartment boundaries within somites. |
| GO:0001756 somitogenesis | IMP PMID:15170697 Dll3 pudgy mutation differentially disrupts dynamic expressi... | ACCEPT | Summary: Dll3 is expressed in the PSM and Dll3 loss (pudgy, targeted null) disrupts somite formation, segmentation-clock gene expression and somite polarity. Reason: Somitogenesis is the principal in vivo process requiring Dll3 (core process). Supporting Evidence: PMID:15170697 We report here that the Dll3(pu) mutation has different effects on the expression of cycling (Lfng and Hes7) and stage-specific genes (Hey3 and Mesp2). PMID:9662403 the pu mutation disrupts the proper formation of morphological borders in early somite formation and of rostral-caudal compartment boundaries within somites. |
| GO:0005112 Notch binding | IBA GO_REF:0000033 | ACCEPT | Summary: IBA (Notch-ligand family node; Dll1, Jag1 and other ligands among donors) for Notch binding. Reason: Dll3 does not bind Notch in trans (PMID:16144902), but it associates with Notch1 in cis in the late endocytic compartment (PMID:21147753) and co-immunoprecipitates with Notch2 (PMID:34635817). The term does not specify trans binding, so the inherited function holds, though its mode (cis, intracellular) has diverged from activating ligands. Supporting Evidence: PMID:21147753 Dll3 is not presented on the surface of presomitic mesoderm (PSM) cells in vivo, but instead interacts with Notch1 in the late endocytic compartment. PMID:16144902 Dll3 does not bind to cells expressing any of the four N receptors, and N1 does not bind Dll3-expressing cells. |
| GO:0005112 Notch binding | ISO GO_REF:0000096 | ACCEPT | Summary: Orthology transfer from rat Dll3 (RGD:70953) of Notch binding. Reason: Consistent with cis association of Dll3 with Notch1 and Notch2 (PMID:21147753, PMID:34635817). |
| GO:0005112 Notch binding | NAS PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | ACCEPT | Summary: NAS from the original Dll3 paper, which inferred that Dll3 activates Notch as a conventional Delta ligand. Reason: The activation inference was later refuted (PMID:16144902), but Notch binding per se is supported by later cis-interaction data; the term itself remains valid. Supporting Evidence: PMID:21147753 interacts with Notch1 in the late endocytic compartment |
| GO:0005509 calcium ion binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro EGF-like calcium-binding domain mapping. Reason: Plausible from Ca2+-binding EGF repeats but not experimentally characterised for Dll3; a structural property rather than a functional role. |
| GO:0005515 protein binding | IPI PMID:16000382 Mind bomb 1 is essential for generating functional Notch lig... | MODIFY | Summary: Mib1 (Q80SY4) interacts with Dll3 and with all other mouse Notch ligands. Reason: Generic protein binding is uninformative. The interaction is with the E3 ubiquitin ligase Mib1, for which Dll3 is a substrate/client, so ubiquitin protein ligase binding is the informative term. Proposed replacements: ubiquitin protein ligase binding Supporting Evidence: PMID:16000382 Mib1 interacts with and regulates all of the Notch ligands, jagged 1 and jagged 2, as well as Dll1, Dll3 and Dll4. |
| GO:0005515 protein binding | IPI PMID:34635817 Gm364 coordinates MIB2/DLL3/Notch2 to regulate female fertil... | MODIFY | Summary: Two IPI rows from the Gm364 oocyte study, with Notch2 (O35516) and Gm364 (A2AFI6) as interactors (co-IP). Reason: Generic protein binding is uninformative. The Notch2 interaction is captured more informatively as Notch binding; the Gm364 interaction does not support any specific molecular function and on its own would be removed. Proposed replacements: Notch binding Supporting Evidence: PMID:34635817 P Co-IP and blot showed that Notch2 interacts with DLL3. PMID:34635817 O Co-IP and blot showed that Gm364 interacts with DLL3. |
| GO:0005515 protein binding | IPI PMID:34635817 Gm364 coordinates MIB2/DLL3/Notch2 to regulate female fertil... | MODIFY | Summary: Two IPI rows from the Gm364 oocyte study, with Notch2 (O35516) and Gm364 (A2AFI6) as interactors (co-IP). Reason: Generic protein binding is uninformative. The Notch2 interaction is captured more informatively as Notch binding; the Gm364 interaction does not support any specific molecular function and on its own would be removed. Proposed replacements: Notch binding Supporting Evidence: PMID:34635817 P Co-IP and blot showed that Notch2 interacts with DLL3. PMID:34635817 O Co-IP and blot showed that Gm364 interacts with DLL3. |
| GO:0005886 plasma membrane | IBA GO_REF:0000033 | KEEP AS NON CORE | Summary: IBA (PANTHER:PTN001170801) plasma membrane, inherited from cell-surface Notch ligands (Dll1, Jag). Reason: Dll3 is a transmembrane protein and has some evidence of surface/membrane localisation (oocyte), but endogenous PSM DLL3 is mainly intracellular (Golgi, late endosome), a divergence from the activating ligands in the node. Supporting Evidence: PMID:17664336 In contrast to endogenous DLL1 located on the surface of presomitic mesoderm cells, we find endogenous DLL3 predominantly in the Golgi apparatus. |
| GO:0005886 plasma membrane | IDA PMID:34635817 Gm364 coordinates MIB2/DLL3/Notch2 to regulate female fertil... | KEEP AS NON CORE | Summary: Immuno-EM detected DLL3 at the oocyte membrane in close proximity to MIB2. Reason: Experimental and accepted for the oocyte context, but in the presomitic mesoderm endogenous DLL3 is predominantly in the Golgi and late endocytic compartments rather than at the cell surface; the plasma membrane is not where its principal function is executed. Supporting Evidence: PMID:34635817 U Immuno-EM showed that MIB2 and DLL3 localized at the oocyte membrane and were close (<20 nm) to each other. |
| GO:0007219 Notch signaling pathway | IBA GO_REF:0000033 | MODIFY | Summary: IBA (PANTHER:PTN001170801) Notch signaling pathway, inherited from activating DSL ligands (Dll1, DLL1 human among donors). Reason: Dll3 has diverged functionally from activating DSL ligands: it neither binds Notch in trans nor activates signalling, and acts only as a cell-autonomous inhibitor. Its relationship to the pathway is captured precisely by negative regulation of Notch signaling pathway (also inherited from the same node), not by participation in the signalling pathway itself. Propagation Review Root cause: PROPAGATION BAD Failure modes: FUNCTIONAL DIVERGENCE Sources checked: PANTHER:PTN001170801 SUPPORTS SOURCE BUT NOT TARGET Node covers activating Notch ligands; Dll3 is a divergent non-activating paralog with cis-inhibitory function only. Proposed replacements: negative regulation of Notch signaling pathway Supporting Evidence: PMID:16144902 In contrast to other DSL ligands, we show that Dll3 does not activate N signaling in multiple assays. |
| GO:0007219 Notch signaling pathway | IEA GO_REF:0000002 | MODIFY | Summary: InterPro (Notch_ligand_N, IPR011651) mapping to Notch signaling pathway. Reason: Domain-based mapping assumes an activating ligand; for Dll3 the evidence supports negative regulation of Notch signalling instead. Proposed replacements: negative regulation of Notch signaling pathway Supporting Evidence: PMID:16144902 Therefore, Dll3 functions not as an activator as previously reported but rather as a dedicated inhibitor of N signaling. |
| GO:0007219 Notch signaling pathway | NAS PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | MODIFY | Summary: NAS from the original Dll3 paper that inferred Dll3 activates Notch. Reason: The activation inference was refuted by direct assays; Dll3 is a dedicated cell-autonomous inhibitor of Notch signalling. Proposed replacements: negative regulation of Notch signaling pathway Supporting Evidence: PMID:16144902 Therefore, Dll3 functions not as an activator as previously reported but rather as a dedicated inhibitor of N signaling. |
| GO:0007386 compartment pattern specification | IMP PMID:12001066 Segmentation defects of Notch pathway mutants and absence of... | KEEP AS NON CORE | Summary: Dll3 mutants show segmentation defects including disrupted somite rostro-caudal compartments. Reason: Somite compartment (rostral/caudal) patterning is disrupted in Dll3 mutants, a component of its somitogenesis role rather than a separate core function. Supporting Evidence: PMID:9662403 the pu mutation disrupts the proper formation of morphological borders in early somite formation and of rostral-caudal compartment boundaries within somites. PMID:12001066 Lfng-deficient mice and Dll3-deficient mice exhibit very similar defects, and marker analysis suggests that progression of the segmentation clock is disrupted in Dll3 mutants. |
| GO:0007399 nervous system development | NAS PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | MARK AS OVER ANNOTATED | Summary: NAS from expression in postmitotic neurons of the spinal cord. Reason: Expression-based; no nervous system development phenotype is documented for Dll3 mutants in the reviewed literature. Supporting Evidence: PMID:9272948 We confirm that, during neurogenesis in the spinal cord, Dll1 and Dll3 are expressed by postmitotic cells |
| GO:0009888 tissue development | IDA PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | MARK AS OVER ANNOTATED | Summary: IDA from the expression paper linking Dll3 to tissue production from primitive streak and tailbud. Reason: Very general term inferred from expression pattern; the specific supported process is somitogenesis. Supporting Evidence: PMID:9272948 The three major sites of expression implicate Dll3 in somitogenesis and neurogenesis and in the production of tissue from the primitive streak and tailbud. |
| GO:0010008 endosome membrane | TAS Reactome:R-MMU-9793875 | ACCEPT | Summary: Reactome TAS placing Dll3 in endosome membrane (Dll3 binds Notch1 in endosomes; expression events). Reason: Consistent with experimental evidence that Dll3 interacts with Notch1 in the late endocytic compartment. Supporting Evidence: PMID:21147753 interacts with Notch1 in the late endocytic compartment |
| GO:0010008 endosome membrane | TAS Reactome:R-MMU-9793917 | ACCEPT | Summary: Reactome TAS placing Dll3 in endosome membrane (Dll3 binds Notch1 in endosomes; expression events). Reason: Consistent with experimental evidence that Dll3 interacts with Notch1 in the late endocytic compartment. Supporting Evidence: PMID:21147753 interacts with Notch1 in the late endocytic compartment |
| GO:0010008 endosome membrane | TAS Reactome:R-MMU-9825303 | ACCEPT | Summary: Reactome TAS placing Dll3 in endosome membrane (Dll3 binds Notch1 in endosomes; expression events). Reason: Consistent with experimental evidence that Dll3 interacts with Notch1 in the late endocytic compartment. Supporting Evidence: PMID:21147753 interacts with Notch1 in the late endocytic compartment |
| GO:0016020 membrane | IEA GO_REF:0000120 | ACCEPT | Summary: Membrane (type I transmembrane protein), from InterPro/UniProt subcellular location. Reason: Correct; Dll3 is a single-pass membrane protein of Golgi/endosomal membranes. |
| GO:0045746 negative regulation of Notch signaling pathway | IBA GO_REF:0000033 | ACCEPT | Summary: Negative regulation of Notch signaling pathway (IBA from PANTHER:PTN001170801; ISO from rat Dll3). Dll3 cell-autonomously inhibits Notch signalling induced by other DSL ligands. Reason: This is the core biological role of Dll3 (cis-inhibition of Notch). Supporting Evidence: file:mouse/Dll3/Dll3-deep-research-falcon.md Dll3 serves primarily as a **cell-autonomous cis-inhibitor** of Notch signaling PMID:16144902 Although Dll3 did not bind or activate N when presented in trans, it cell autonomously inhibited N signaling that was induced by other DSL ligands in CSL gene reporter, Xenopus laevis primary neurogenesis, and mouse embryonic neural progenitor differentiation assays. PMID:21147753 Dll3 targeting newly synthesized Notch1 for lysosomal degradation prior to post-translational processing and cell surface presentation of the receptor. |
| GO:0045746 negative regulation of Notch signaling pathway | ISO GO_REF:0000096 | ACCEPT | Summary: Negative regulation of Notch signaling pathway (IBA from PANTHER:PTN001170801; ISO from rat Dll3). Dll3 cell-autonomously inhibits Notch signalling induced by other DSL ligands. Reason: This is the core biological role of Dll3 (cis-inhibition of Notch). Supporting Evidence: file:mouse/Dll3/Dll3-deep-research-falcon.md Dll3 serves primarily as a **cell-autonomous cis-inhibitor** of Notch signaling PMID:16144902 Although Dll3 did not bind or activate N when presented in trans, it cell autonomously inhibited N signaling that was induced by other DSL ligands in CSL gene reporter, Xenopus laevis primary neurogenesis, and mouse embryonic neural progenitor differentiation assays. PMID:21147753 Dll3 targeting newly synthesized Notch1 for lysosomal degradation prior to post-translational processing and cell surface presentation of the receptor. |
| GO:0048339 paraxial mesoderm development | IMP PMID:15170697 Dll3 pudgy mutation differentially disrupts dynamic expressi... | KEEP AS NON CORE | Summary: Dll3 pudgy mutation disrupts cycling and stage-specific gene expression in paraxial mesoderm. Reason: Captured more precisely by somitogenesis. Supporting Evidence: PMID:15170697 We report here that the Dll3(pu) mutation has different effects on the expression of cycling (Lfng and Hes7) and stage-specific genes (Hey3 and Mesp2). |
| GO:0048712 negative regulation of astrocyte differentiation | ISO GO_REF:0000096 | KEEP AS NON CORE | Summary: ISO from rat Dll3. Dll3 misexpression inhibited glial differentiation of mouse neural progenitors, as a Notch antagonist. Reason: Based on overexpression in cultured neural progenitors; a context-specific consequence of Notch antagonism, without in vivo loss-of-function support. Supporting Evidence: PMID:16144902 As an N antagonist, Dll3 promoted Xenopus laevis neurogenesis and inhibited glial differentiation of mouse neural progenitors. |
| GO:0050768 negative regulation of neurogenesis | IDA PMID:9272948 Mouse Dll3: a novel divergent Delta gene which may complemen... | REMOVE | Summary: Dunwoodie et al. 1997 reported that ectopic Dll3 inhibited primary neurogenesis in Xenopus and interpreted Dll3 as an activating ligand. Reason: Contradicted: a later systematic study found that Dll3 does not activate Notch and, as a Notch antagonist, promoted rather than inhibited Xenopus primary neurogenesis. The original conclusion rested on the now-refuted assumption that Dll3 activates Notch, and no in vivo loss-of-function evidence supports negative regulation of neurogenesis. Supporting Evidence: PMID:16144902 As an N antagonist, Dll3 promoted Xenopus laevis neurogenesis and inhibited glial differentiation of mouse neural progenitors. PMID:9272948 We confirm that Dll3 can inhibit primary neurogenesis when ectopically expressed in Xenopus, suggesting that it can activate the Notch receptor |
| GO:0050768 negative regulation of neurogenesis | IEA GO_REF:0000117 | REMOVE | Summary: ARBA machine-learning prediction of negative regulation of neurogenesis. Reason: Rule-based transfer from activating Delta ligands; for Dll3, overexpression data point in the opposite direction (promotion of neurogenesis as a Notch antagonist) and there is no loss-of-function support. Supporting Evidence: PMID:16144902 As an N antagonist, Dll3 promoted Xenopus laevis neurogenesis |
| GO:0050769 positive regulation of neurogenesis | ISO GO_REF:0000096 | KEEP AS NON CORE | Summary: ISO from rat Dll3; Dll3 as Notch antagonist promoted Xenopus neurogenesis. Reason: Consistent with Dll3 being a Notch inhibitor, but based on heterologous overexpression; not a demonstrated in vivo role. Supporting Evidence: PMID:16144902 As an N antagonist, Dll3 promoted Xenopus laevis neurogenesis |
| GO:0005794 Golgi apparatus | IDA PMID:17664336 Divergent functions and distinct localization of the Notch l... | NEW | Summary: Endogenous DLL3 in presomitic mesoderm is predominantly located in the Golgi apparatus. Reason: Direct localisation of the endogenous protein in vivo; this is the main site of Dll3 in PSM cells and is missing from GOA. Supporting Evidence: PMID:17664336 In contrast to endogenous DLL1 located on the surface of presomitic mesoderm cells, we find endogenous DLL3 predominantly in the Golgi apparatus. |
| GO:0005770 late endosome | IDA PMID:21147753 Notch inhibition by the ligand DELTA-LIKE 3 defines the mech... | NEW | Summary: Dll3 interacts with Notch1 in the late endocytic compartment in PSM cells. Reason: Location where Dll3-Notch1 cis-interaction occurs; more specific than endosome membrane. Supporting Evidence: PMID:21147753 Dll3 is not presented on the surface of presomitic mesoderm (PSM) cells in vivo, but instead interacts with Notch1 in the late endocytic compartment. |
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Download this section (compressed HTML)Q: Is Dll3-mediated cis-inhibition of Notch1 in the PSM physiologically significant, given that Geffers et al. (2007) found no evidence for DLL3 antagonism of DLL1 or repression of Notch activity in vivo, whereas Ladi et al. (2005) and Chapman et al. (2011) report cis-inhibition?
Q: Does GO need a cis-specific Notch-binding or "Notch receptor cis-inhibitor activity" term to distinguish non-activating ligands such as Dll3 from trans-activating DSL ligands?
Experiment: Measure Notch1 processing and surface levels in Dll3-null versus wild-type PSM with an endogenous-tagged Notch1, to test whether Dll3 routes nascent Notch1 to lysosomes in vivo.
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