Zebrafish embryos exposed to the model hedgehog pathway inhibitor cyclopamine, as well as to the potential hedghog pathway disrupters SANT-2 and GANT-61.
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE29357 | GEO | 2011/10/01
SECONDARY ACCESSION(S): PRJNA140187
REPOSITORIES: GEO
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Project description:The hedgehog (Hh) signaling pathway is important for various developmental processes during embryogenesis and for homeostasis of adult tissue in vertebrates. Aberrant signaling results in severe birth defects and malignancies, respectively. The small molecules SANT-2 and GANT-61 were identified as potential Hh-pathway inhibitors in a reporter gene assay (Shh-light II cells). We aimed to analyze the effects of these compounds on the gene expression level in zebrafish embryos in comparison with the well known Hh-pathway inhibitor cyclopamine. Zebrafish embryos were treated with Ethanol (0,1%) (control), 10 µM Cyclopamine (C), 10 µM SANT-2 (S), and 10 µM GANT-61 (G), for 24 h (0.5 - 24 hpf), respectively; all in 5 independent replicates with 100 zebrafish embryos per treatment. Total RNA was extracted and processed as recommended by Agilent. One color arrays were used and data is given as normalized log2 of the gprocessed signals.
2011-09-30 | E-GEOD-29357 | biostudies-arrayexpress
Project description:Analysis of gene expression levels in response to inhibition of Hh signaling in HT-29 cells using a cDNA microarray technique. Microarray analyses revealed that different genes after treatment with both cyclopamine (an antagonist of SMO) and GANT61 (a specific, small-molecular molecule inhibitor of Glis).
2021-06-22 | GSE116218 | GEO
Project description:T helper 17 (Th17) cells play a key role in barrier protection against fungal and bacterial pathogens but are also pathological drivers of many inflammatory diseases. We find that Hh signaling inhibition through Smoothened inhibitor cyclopamine selectively inhibits Th17 lineage differentiation but not the development of Th1, Th2, or iTreg CD4+ Th cells. In these experiments we analysed the transcriptomes of Th17 cells that had been polarized in the presence of cyclopamine or carrier control to profile the global transcriptome of Th17 cells in which the Hh pathway had been blocked. The aim of this analysis was to identify the target genes downstream of Smo that may be important for Th17 polarisation.
2022-06-13 | GSE205848 | GEO
Project description:Physiologically, trophoblast progenitor cells differentiate into placental villous cytotrophoblast cells (CTBs). CTBs either differentiate into invasive lineage to yields extravillous cytotrophoblast cells (EVTs), or undergo cell fusion lineage to yields syncytiotrophoblast cells (STBs)，Sonic hedgehog (Shh) together with indian hedgehog (Ihh) and desert hedgehog (Dhh) consist of ligand of hedgehog signaling pathway, which plays pivotal roles in regulating cell proliferation, cell differentiation, organogenesis and development, even involving in tumorigenesis and progression. previous study had summarized and indicatedthat hedgehog proteins played important roles in regulating hematopoiesis, vasculogenesis and angiogenesis during embryogenesis and development. Herein, we investigate the effect of the Sonic Hedgehog morphogen inhibitor Cyclopamine on JAR cells
2019-12-31 | GSE130367 | GEO
Project description:Background. Primary cilia (PC) are solitary antennae present at the cell surface. These non-motile cilia play an important role in organ development and tissue homeostasis through the transduction of the Hedgehog (Hh) signaling pathway. We recently revealed the presence of PC in the epithelium of the developing epididymis, an organ of the male reproductive system whose dysfunction triggers male infertility. Acknowledging that systemic blockade of the Hh pathway trigger epididymal dysfunctions in vivo, our main goals were 1) to portray the epididymal Hh environment, 2) to determine the direct responsiveness of epididymal epithelial cells to Hh, and 3) to define the contribution of PC to the transduction of this pathway. Results. The Hh ligands Indian and Sonic hedgehog (Ihh and Shh) were respectively located in principal and clear cells of the mouse epididymis by immunofluorescent staining. The propensity of epididymal principal cells to respond to Hh signaling was assessed on immortalized epididymal DC2 cells by western-blot, confocal imaging and 3D-reconstruction. Our results indicate that epididymal principal cells secrete Ihh and expose PC that co-localize with the conventional acetylated tubulin/Arl13b ciliary markers, as well as with GLI3 Hh signaling factor. Gene expression microarray profiling indicated that the expression of 43 and 248 genes was respectively and significantly modified following pharmacological treatment of DC2 cells with the Hh agonist SAG (250 nM) or the Hh antagonist cyclopamine (20 µM) compared with the control. Among Hh target genes identified, 6.7 % presented perfect matches for GLI-transcription factor consensus sequences, and the majority belonged to interferon-dependent immune response and lipocalin 2 pathways. Finally, the contribution of epididymal PC to the transduction of canonical Hh pathway was validated by ciliobrevinD treatment, which induced a significant decrease of PC length and the expressional reduction of Hh signalling targets. Conclusions. All together our data indicate that PC from epithelial principal cells regulate gene expression profile through a possible autocrine Hh signaling. This provides new hypotheses regarding the potential contribution of PC and Hh signaling in intercellular cross-talk and immunological regulation of the epididymis.
2020-04-17 | GSE143829 | GEO
Project description:Aberrant activation of Hedgehog pathway is responsible for initiation and maintenance of various cancers, including medulloblastoma (MB), basal cell carcinoma (BCC), and other solid and hematological tumors. Therefore, targeting Hh pathway represents promising therapeutic prospects for Hh-driven cancers. In recent years, tremendous efforts have been dedicated to the discovery of Hh pathway inhibitor. While the majority of Hh pathway inhibitors target the upstream membrane protein Smoothened (SMO). Here, we performed Next Generation Sequencing to reveal the target genes of Hh pathway by treating mouse SHH-subtype medulloblastoma cells (SmoWT) with SMO inhibitor (GDC0449) or DMSO.
2021-07-08 | GSE171689 | GEO
Project description:Introduction and objectives: Proper In utero Wolffian duct (WD) development requires androgens and is essential to epididymis formation and male fertility. However, non-hormonal factors that control WD homeostasis and development remain largely unknown. In this study, we investigated the contribution of Hedgehog signaling pathway to Wolffian duct development by combining pharmacological approaches on organotypic cultures of WD with microarray profiling. Methods: WD were collected from embryos isolated from 16.5 days post-coitum pregnant mice and cultured in air-liquid condition at 37oC during 72 hours in the presence of either: 1) Cyclopamine (25 µM): Hh inhibitor; 2) Smoothened agonist (0.5 µM ; SAG): Hh activator; 3) Culture media (control). WD pictures were taken and analyzed on ImageJ. At the end of the culture, WD were snap-frozen to perform transcriptomic microarray analyses. Three to four biological replicates per condition, i.e. control (n=3), SAG (n=4) and cyclopamine (n=3), were used for microarray analyses. The quality of the RNA was determined using an Agilent BioAnalyzer (Agilent Technologies, Santa Clara, CA, USA) and displayed a RIN ranging from 9.10 to 9.60 out of 10 for the different samples. Microarray analyses were carried out on Affymetrix Mouse Clariom S arrays (Thermofisher) according to the Affymetrix standard protocol. In brief, 100 ng total RNA samples were labeled using the GeneChip® WT Plus Reagent Kit protocol and hybridized to the arrays as described by the manufacturer (Affymetrix, Thermofisher). The cRNA hybridization cocktail was incubated overnight at 45°C while rotating in a hybridization oven. After a 16-h hybridization period, the cocktail was removed and the arrays were washed and stained in an Affymetrix GeneChip fluidics station 450, according to the Affymetrix-recommended protocol. The arrays were scanned using the Affymetrix GCS 3000 7G and Gene-Chip Command Console Software (AGCC) (Affymetrix, Thermofisher) to produce the probe cell intensity data (CEL). The imaged data were then analyzed using the Affymetrix Expression Console Software to perform the quality control, background subtraction and normalization of probe set intensities using Robust Multiarray Analysis (RMA). Microarray processing was performed by the Gene Expression Core facility of the Genomic platform of the Centre Hospitalier Universitaire de Quebec Research Center. The microarray CEL files were imported and analyzed with Transcriptome Analysis Console (TAC) Software 4.0.2 (Applied Biosystems), and submitted to RMA normalization. Samples were included within three different treatment groups (control, SAG and cyclopamine) and compared by Analysis of Variance. Principal Component Analysis and heat maps were performed with Partek Pathway (Partek Incorporated). Biological pathways analyses were performed by using 5 distinct algorithms (i.e. DAVID, STRING, GOrilla, Metascape, GSEA and BLAST2GO). Conclusion: The data made available on GEO repository provide new insights regarding the mechanisms that control epididymis morphogenesis and male fertility. Financially supported by a CIHR grant to CB.
2022-02-21 | GSE145816 | GEO
Project description:Basal cell carcinomas (BCCs) rely on Hedgehog (HH) pathway growth signal amplification by the microtubule-based organelle, the primary cilium. Despite naïve tumors responsiveness to Smoothened inhibitors (Smoi), resistance in advanced tumors remains frequent. While the resistant BCCs usually maintain HH pathway activation, squamous cell carcinomas with Ras/MAPK pathway activation also arise, with the molecular basis of tumor type and pathway selection still obscure. Here we identify the primary cilium as a critical determinant controlling tumor pathway switching. Strikingly, Smoi-resistant BCCs possess an increased mutational load in ciliome genes, resulting in reduced primary cilia and HH pathway activation compared to naive or Gorlin patient BCCs. Gene set enrichment analysis of resistant BCCs with a low HH pathway signature reveals increased Ras/MAPK pathway activation. Tissue analysis confirms an inverse relationship between primary cilia presence and Ras/MAPK activation, and primary cilia removal in BCCs potentiates Ras/MAPK pathway activation. Moreover, activating Ras in HH-responsive cell lines confers resistance to both canonical (vismodegib) and non-canonical (aPKC and MRTF inhibitors) HH pathway inhibitors, while conferring sensitivity to MAPK inhibitors. Our results provide new insights into BCC treatment and identify the primary cilium as an important lineage gatekeeper, preventing HH to Ras/MAPK pathway switching.
2018-10-10 | GSE120954 | GEO
Project description:Sonic Hedgehog inhibitor cyclopamine effect on JAR cells
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Project description:Target deconvolution of small molecule hits from phenotypic screens presents a major challenge. Illustrative of these are the many screens that have been conducted to find inhibitors for the Hedgehog (Hh) signaling pathway – a major developmental pathway with many implications in health and disease - with many hits but very few identified cellular targets. We here present a strategy for target identification based on Proteolysis-Targeting Chimeras (PROTACs), combined with label-free quantitative proteomics. We developed a PROTAC based on the downstream Hedgehog Pathway Inhibitor-1 (HPI-1), a phenotypic screen hit with unknown cellular target. Using our Hedgehog Pathway PROTAC (HPP) we identified and validated BET bromodomains to be the cellular targets of HPI-1. Furthermore, we found that HPP-9 has a unique mechanism of action as a long-acting Hh pathway inhibitor through prolonged BET bromodomain degradation. Collectively, we provide a powerful PROTAC-based approach for target deconvolution, that has answered the longstanding question of the cellular target of HPI-1 and yielded the first PROTAC that acts on the Hh pathway.
2023-07-20 | PXD036539 | Pride