| 2002 |
Genetic epistasis in mice showed that Shh and Gli3 are dispensable for limb skeletal element formation; Shh(-/-) Gli3(-/-) double mutants have distally complete but polydactylous limbs lacking normal digit identities. The effects of Shh signaling on skeletal patterning are necessarily mediated through Gli3, by regulating the balance of Gli3 transcriptional activator and repressor activities. |
Genetic double-mutant analysis (Shh-/- Gli3-/- mice), skeletal preparation and phenotypic analysis |
Nature |
High |
12198547
|
| 1998 |
Shh protein applied ectopically to mandibular mesenchyme induced expression of Ptc and Gli1, demonstrating that SHH can act as a direct inducer of these pathway targets. Ectopic Shh within tooth germs caused abnormal epithelial invagination, indicating a role for Shh in epithelial cell proliferation during tooth development. |
Ectopic protein application to explants; in vivo organ culture; mutant analysis (Gli2/Gli3 double mutants) |
Development (Cambridge, England) |
High |
9655803
|
| 2003 |
Using mutant analysis and in vitro explant assays, Gli2 and Gli3 were shown to be required for Shh-dependent sclerotome induction; somitic mesoderm from Gli2(-/-)Gli3(-/-) embryos cannot activate sclerotomal genes in response to exogenous Shh. Additionally, Gli2 was shown to have a repressor function and Gli3 an activator function in the somite, and each Gli preferentially activates a distinct subset of Shh target genes. |
Genetic mutant analysis, in vitro explant assays, adenoviral Gli overexpression in presomitic mesoderm explants |
Development (Cambridge, England) |
High |
14602680
|
| 2000 |
Cholesterol is important for SHH biogenesis, and teratogens that induce holoprosencephaly (cyclopamine, cholesterol synthesis inhibitors) affect Shh signal transduction in responding cells rather than Shh biogenesis itself. The structural similarity of the Shh receptor Patched (Ptc) to the Niemann-Pick C1 protein (involved in vesicular cholesterol trafficking) implicates cholesterol in Shh signal transduction. |
Pharmacological inhibitor studies (cyclopamine, cholesterol synthesis inhibitors); review of mechanistic data on Shh processing and pathway activation |
Cellular and molecular life sciences : CMLS |
Medium |
11130177
|
| 2006 |
FGF9 and SHH signaling coordinate lung mesenchymal development through distinct sub-mesothelial and sub-epithelial compartments. FGF9 signals from the epithelium to sub-epithelial mesenchyme to maintain SHH signaling, which regulates cell proliferation, survival and mesenchymal-to-epithelial signaling. FGF9 alone can only partially rescue vascular defects caused by SHH loss, and SHH cannot rescue FGF9-null vascular phenotypes, indicating they regulate distinct aspects of development. |
Loss-of-function and inducible gain-of-function mouse models (Fgf9 KO, Shh signaling loss), phenotypic and gene expression analysis |
Development (Cambridge, England) |
High |
16540513
|
| 2007 |
FGF9 and SHH signaling to lung mesenchyme (but not to endothelial cells) are each necessary and together sufficient for distal pulmonary capillary development, acting by regulating Vegfa expression in lung mesenchyme. VEGF signaling is required downstream of FGF9-mediated blood vessel formation. |
Gain- and loss-of-function genetics in mice, conditional deletion, Vegfa expression analysis, vascular morphometry |
Development (Cambridge, England) |
High |
17881491
|
| 2009 |
BMP activity negatively regulates Shh transcription in the limb bud, forming a BMP-Shh negative-feedback loop that confines Shh expression to the ZPA. BMP-dependent downregulation of Shh is achieved by interfering with FGF and Wnt signaling activities that maintain Shh expression. FGF induction of Shh requires protein synthesis and is mediated by the ERK1/2 MAPK pathway. |
In vivo limb bud manipulation, pharmacological inhibitors (ERK1/2 MAPK), gene expression analysis, gain- and loss-of-function experiments |
Development (Cambridge, England) |
High |
19855020
|
| 2011 |
Lhx6 and Lhx8 transcription factors coexpressed in early-born MGE neurons are required to induce neuronal Shh expression. Shh function in early-born MGE neurons feeds forward to promote SHH signaling in the overlying progenitor zone, regulating Lhx6, Lhx8, and Nkx2-1 expression and production of late-born somatostatin+ and parvalbumin+ cortical interneurons. |
Genetic conditional deletion of Shh in MGE mantle zone, gene expression analysis, interneuron counting |
Neuron |
High |
21658586
|
| 2010 |
Foxa2 positively regulates Shh expression in multiple tissues, but in the midbrain Foxa1 and Foxa2 attenuate Shh signaling by directly inhibiting expression of its intracellular transducer Gli2 at the transcriptional level. ChIP experiments showed that Foxa2 binds to genomic regions of Gli2. |
Conditional KO of Foxa2 in midbrain (Wnt1cre;Foxa2flox/flox), gain-of-function studies in mice, chromatin immunoprecipitation (ChIP) |
Mechanisms of development |
High |
21093585
|
| 2015 |
The Shh gradient amplitude in the mouse neural tube increases over time, but Gli transcriptional effector activity initially increases then decreases (adaptation). Computational and experimental analysis identified three contributing mechanisms: transcriptional upregulation of inhibitory receptor Ptch1, transcriptional downregulation of Gli, and differential stability of active vs. inactive Gli isoforms. Gli2 protein expression is downregulated during neural tube patterning, and adaptation continues when the pathway is stimulated downstream of Ptch1. |
Quantitative imaging of Shh gradient in developing mouse neural tube, computational modeling, Gli2 protein expression analysis, NIH3T3 cell culture experiments |
Nature communications |
High |
25833741
|
| 2014 |
Ptch2 mediates the Shh response in Ptch1-/- cells. The Shh response in Ptch1(-/-) cells is ligand-dependent and can be inhibited by Shh-blocking antibody 5E1. Ptch1(-/-);Ptch2(-/-) double KO cells cannot further activate the Shh response, demonstrating that Ptch2 mediates Shh signaling in the absence of Ptch1. Expression of dominant-negative Ptch2 in developing chick neural tube caused activation of the Shh response, indicating Ptch2 suppresses Shh signaling at early developmental stages. |
Ptch1/Ptch2 double KO cells, Shh-blocking antibody (5E1), dominant-negative Ptch constructs in chick neural tube electroporation, cell migration assays |
Development (Cambridge, England) |
High |
25085974
|
| 2014 |
Shh-induced activation of Smoothened (Smo) drastically increases Hhip (Hedgehog-interacting protein) internalization and degradation cell-autonomously. While Hhip can leave its site of synthesis to inhibit Shh non-cell-autonomously, it cannot cell-autonomously inhibit the consequences of Smo activation. This provides a mechanism by which Shh activates pathway response while negating cell-autonomous effects of Hhip, yet Hhip retains non-cell-autonomous inhibitory capacity. |
Hhip overexpression and localization assays, Smo activation studies, internalization and degradation assays |
Nature communications |
Medium |
25215859
|
| 2014 |
Boc (a Shh-binding protein) associates with the Shh receptor Ptch1 to mediate Shh signaling. Boc, through elevated Shh signaling, promotes high levels of DNA damage mediated by CyclinD1. High DNA damage in the presence of Boc increases the incidence of Ptch1 loss of heterozygosity, driving progression from early to advanced medulloblastoma. |
Boc genetic inactivation in mice, medulloblastoma progression analysis, CyclinD1 mechanistic studies, DNA damage assays |
Developmental cell |
Medium |
25263791
|
| 2015 |
Eya1 phosphatase, acting together with the DNA-binding protein Six1, promotes gene induction in response to Shh by regulating Gli transcriptional activators. Eya1 was identified via shRNA screen of the phosphatome and is required for Shh-dependent hindbrain growth; catalytically active Eya1 (its phosphatase activity) is necessary for this function. |
shRNA screen of phosphatome, loss-of-function and gain-of-function assays, catalytic mutant analysis, in vivo genetic analysis of hindbrain development |
Developmental cell |
High |
25816987
|
| 2018 |
Shh-mediated axon guidance of commissural neurons requires Dock3/4 GEFs and their binding partners ELMO1/2. Mechanistically, Dock and ELMO interact with Boc (the Shh receptor), and this interaction is reduced upon Shh stimulation. Shh stimulation translocates ELMO to the growth cone periphery and activates Rac1, identifying Dock/ELMO as an effector complex of non-canonical Shh signaling for growth cone turning. |
shRNA knockdown in vitro axon turning assays, in vivo commissural axon guidance analysis, co-immunoprecipitation of Dock/ELMO with Boc, Rac1 activation assays, subcellular localization imaging |
Developmental cell |
High |
30078728
|
| 2007 |
Protease nexin 1 (PN-1/SERPINE2) interacts with LRP (low-density lipoprotein receptor-related proteins) to antagonize SHH-induced CGNP proliferation and inhibit GLI1 transcriptional activity. PN-1 binding to LRPs interferes with SHH-induced cyclin D1 expression. CGNPs from Pn-1-deficient mice show enhanced basal proliferation due to overactivation of the SHH pathway. |
PN-1 knockout mouse analysis, proliferation assays, GLI1 activity assays, cyclin D1 expression analysis, binding studies with LRP |
Development (Cambridge, England) |
High |
17409116
|
| 2019 |
Highly recurrent U1 snRNA hotspot mutations (r.3A>G) in ~50% of SHH medulloblastomas occur in the 5' splice-site binding region and cause disrupted RNA splicing with excess 5' cryptic splicing events. Mutant U1 snRNA-mediated alternative splicing inactivates tumor-suppressor PTCH1 and activates oncogenes GLI2 and CCND2, identifying a non-canonical mechanism of SHH pathway activation. |
Whole-genome/transcriptome sequencing of 250 SHH medulloblastomas, splicing analysis, functional characterization of U1 snRNA mutations |
Nature |
High |
31664194
|
| 2016 |
In developing hair buds, SHH signaling is differentially distributed between asymmetric daughter cells: displaced WNT-low suprabasal daughters become stem cells that respond to paracrine SHH and symmetrically expand, while basal daughters express but do not respond to SHH. This WNT-SHH antagonism specifies and expands stem cells prior to niche formation. |
Live imaging, immunofluorescence, genetics, cell-cycle analyses, in utero lentiviral transduction, lineage tracing |
Cell |
High |
26771489
|
| 2021 |
YAP transcription activity, activated downstream of GNAS loss, directly drives Shh expression. Secreted SHH in turn induces YAP activation, Shh expression, and osteoblast differentiation in surrounding wild-type cells, forming a self-amplifying YAP-SHH loop that is necessary and sufficient for heterotopic ossification expansion. Genetic or pharmacological inhibition of either YAP or SHH abolished HO. |
Mouse models of POH (Gnas KO) and FOP, genetic ablation and pharmacological inhibition of YAP and SHH, gain-of-function experiments, gene expression analysis |
Science translational medicine |
High |
34162750
|
| 2022 |
O-GlcNAc transferase (OGT) regulates granule neuron precursor neurogenesis by activating the Shh signaling pathway via O-GlcNAcylation at S355 of Gli2. This modification promotes Gli2 deacetylation and transcriptional activity through dissociation from p300 (a histone acetyltransferase). OGT inhibition improves survival in a medulloblastoma mouse model. |
OGT conditional KO, O-GlcNAcylation site mapping (S355 of Gli2), Gli2-p300 co-immunoprecipitation, acetylation/deacetylation assays, medulloblastoma mouse model |
Proceedings of the National Academy of Sciences of the United States of America |
High |
35969743
|
| 2022 |
Wnt signaling directly regulates epithelial expression of Sonic Hedgehog (SHH), which in turn acts on mesenchymal cells to drive villi formation during small intestine morphogenesis. Subepithelial mesenchymal cell gradients supporting Wnt signaling regulate epithelial SHH expression as part of a mesenchymal-epithelial crosstalk. |
Single-cell analysis, in vitro organoid culture, genetic manipulation, in situ hybridization, gene expression analysis |
Nature communications |
Medium |
35132078
|
| 2019 |
A prechordal enhancer (SBE7) was identified that directs Shh expression in both the prechordal plate and ventral midline of the forebrain. Deletion of SBE7 from the mouse genome markedly downregulated Shh in the rostral axial mesoderm and ventral forebrain/hypothalamus, causing craniofacial abnormality resembling human holoprosencephaly. Prechordal SHH signaling triggers secondary Shh induction in the forebrain, which then directs neuronal differentiation. |
Enhancer identification, targeted deletion from mouse genome, in vivo reporter assays, gene expression analysis |
Proceedings of the National Academy of Sciences of the United States of America |
High |
31685615
|
| 2019 |
Abrogating constitutive transcription over the ZRS enhancer shifts Shh-ZRS contacts and moderately reduces Shh transcription. Deletion of CTCF binding sites around the ZRS results in loss of the preformed Shh-ZRS interaction and 50% decrease in Shh expression but no detectable phenotype, indicating an additional CTCF-independent mechanism. Combining CTCF binding site loss with a hypomorphic ZRS allele causes severe Shh loss of function and digit agenesis. |
CTCF binding site deletion, hypomorphic ZRS allele, chromosome conformation capture, in vivo mouse genetics |
Proceedings of the National Academy of Sciences of the United States of America |
High |
31147463
|
| 2016 |
Foxf2 (downstream of Shh signaling) is required in neural crest-derived palatal mesenchyme for palatogenesis; Foxf1 and Foxf2 together repress Fgf18 expression in the mesenchyme, which is necessary to maintain Shh expression in the palatal epithelium. Addition of exogenous Fgf18 protein to cultured palatal explants directly inhibited Shh expression, establishing a Shh-Foxf-Fgf18-Shh molecular circuit. |
Cre/loxP tissue-specific conditional KO, RNA-seq, whole-mount in situ hybridization, palatal explant culture with exogenous FGF18 protein |
PLoS genetics |
High |
26745863
|
| 2023 |
YAP, acting as a mechanosensor, is activated by a gradient of mechanical stress and tissue stiffness in the notochord and ventral neural tube. YAP activation induces FoxA2 and Shh expression; Hedgehog signaling activation rescues neural tube patterning defects caused by Yap deficiency (but not notochord formation), establishing that mechanotransduction via Yap acts in a feedforward mechanism to activate Shh expression for floor plate induction. |
Yap conditional KO mice, gain-of-function experiments, mechanical stress measurement, tissue stiffness analysis, Hedgehog signaling rescue experiments |
Science advances |
High |
37315133
|
| 2022 |
ETV2 acts as a pioneer transcription factor that initiates Shh expression by changing chromatin status at the ZRS limb enhancer. Etv2 expression precedes Shh in limb buds; Etv2 inactivation prevents ZRS chromatin opening and abolishes Shh expression. Etv2 overexpression causes nucleosomal displacement at ZRS, ectopic Shh expression, and polydactyly. ETV2 is also antagonized by ETV4/5 repressors, and known human polydactyl mutations introduce novel ETV2 binding sites in ZRS. |
Etv2 conditional KO, gain-of-function overexpression, ATAC-seq chromatin accessibility, nucleosome displacement assays, luciferase reporter assays for ETV2 binding sites |
Nature communications |
High |
35864091
|
| 2013 |
Notochord/floor plate-derived Shh regulates mesonephric tubule number and position through indirect effects on the paraxial mesoderm (rather than direct regulation). Mesonephros-specific Shh ablation showed that locally-expressed Shh is not required for mesonephric development. Stage-specific ablation and lineage analysis demonstrated that midline-derived Shh regulates nephrogenic gene expression indirectly via the paraxial mesoderm. |
Shh conditional KO (Hoxb7-Cre, Sall1CreERT2, ShhCreERT2), stage-specific ablation, lineage analysis of Hh-responsive cells, gene expression analysis |
Developmental biology |
High |
24370450
|
| 2020 |
SHH binding to PTCH not only activates the canonical pathway but also blocks PTCH-induced apoptosis (PTCH functions as a dependence receptor that triggers apoptosis in the absence of SHH). Autocrine SHH interference in colon, pancreatic, and lung cancer cell lines triggered cell death through PTCH proapoptotic signaling without changing canonical pathway activity. In vivo, SHH interference decreased primary tumor growth and metastasis. |
SHH interference (knockdown/blocking) in cancer cell lines, in vivo xenograft models, apoptosis assays, canonical pathway activity measurement |
Cancer research |
Medium |
32060146
|
| 2020 |
Downregulation of Shh signaling in the hair matrix is a critical early event in chemotherapy-induced alopecia (CIA). Inhibition of Shh signaling recapitulated key morphological features of CIA, and recombinant Shh protein partially rescued hair loss. Phosphoproteomics identified MAPK pathway activation as a key upstream event that controls Shh downregulation. Shh signaling is an evolutionarily conserved target in CIA pathobiology. |
Mouse model of CIA, recombinant Shh protein rescue, Shh signaling inhibition, phosphoproteomics, human hair follicle organ culture |
The Journal of investigative dermatology |
Medium |
32682910
|
| 2020 |
In developing zebrafish tuberal/anterior hypothalamus, Shh acts as an on-off switch for the homeodomain transcription factor Rx3. Shh coordinates progenitor cell selection and behavior in the tuberal/anterior hypothalamus; in absence of Shh, the shh+ anterior recess does not form and resident differentiated cell types fail to develop. |
rx3 chk mutant/morphant zebrafish, Shh signaling manipulation, EdU pulse-chase, gene expression analysis |
Development (Cambridge, England) |
Medium |
27317806
|
| 2019 |
Genomic analysis mapped Shh-responsive genes in the otic vesicle using Smo loss-of-function and Shh gain-of-function mouse mutants. Gli2 ChIP-seq combined with ATAC-seq identified inner ear enhancers near Shh-responsive genes, revealing Shh-dependent transcriptional networks controlling cochlear duct morphogenesis. |
Comparative transcriptomics of Smo KO and Shh gain-of-function mutants, ATAC-seq, Gli2 ChIP-seq |
Development (Cambridge, England) |
High |
31488567
|