Spatiotemporal regulation of GLI target genes in the mammalian limb bud
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: jordan lewandowski
PROVIDER: E-GEOD-58222 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:GLI proteins convert Sonic hedgehog (Shh) signaling into a transcriptional output in a tissue-specific fashion. The Shh pathway has been extensively studied in the limb bud, where it helps regulate growth through a SHH-FGF feedback loop. However, the transcriptional response is still poorly understood. We addressed this by determining the gene expression patterns of approximately 200 candidate GLI-target genes and identified three discrete SHH-responsive expression domains. GLI-target genes expressed in the three domains are predominately regulated by derepression of GLI3 but have different temporal requirements for SHH. The GLI binding regions associated with these genes harbor both distinct and common DNA motifs. Given the potential for interaction between the SHH and FGF pathways, we also measured the response of GLI-target genes to inhibition of FGF signaling and found the majority were either unaffected or upregulated. These results provide the first characterization of the spatiotemporal response of a large group of GLI-target genes and lay the foundation for a systems-level understanding of the gene regulatory networks underlying SHH-mediated limb patterning.
| S-EPMC4587286 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below.
2015-07-01 | GSE58222 | GEO
Project description:Shh signal mediated by Gli family of transcription factors regulates digit growth and patterning in early limb development. Shh expression in the posterior margin of the limb bud defines the zone of polarizing activity. However, much less is know about downstream targets that mediate Shh signal functions. In this dataset, we include the expression data obtained from dissected anterior and posterior halves of mouse limb bud respectively. These data are used to obtain 889 transcripts that were upregulated 1.3 fold or more in the posterior limb bud, and 1189 transcripts that were enriched in the anterior limb bud at 1.3 fold or more. Two samples were analyzed. We generate pairwise comparisons between anterior and posterior limb tissues. Genes with a fold-change ≥1.3 were selected.
2013-01-06 | E-GEOD-41691 | biostudies-arrayexpress
Project description:Limb buds were dissected from E10.75 mouse embryos and stored in RNAlater reagent (Qiagen), for genotyping. For each replicate, RNA was isolated from pools of 6 limb buds either of wild type or homozygous mutants using RNeasy micro-kit (Qiagen). rRNA was depleted using RiboMinusTM Human/Mouse Transcriptome Isolation Kit (Invitrogen). After cRNA amplification, single or double-stranded cDNA was generated using The GeneChip Whole Transcript Amplified Double-Stranded Target Assay kit (Affymetrix) according to manufacturers instructions. cDNA was fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix) and hybridized to oligonucleotide tiling arrays. The control genomic DNA samples were fragmented with DNAse I. RNA-chip data were computed at the exon level, by averaging the normalized intensities of all probes falling within the exon. As a complement, array data were quantile normalized within cDNA/genomic DNA replicate groups and scaled to medial feature intensity of 10 using TAS software (Affymetrix). For each genomic position, a dataset was generated consisting of all probes mapping within a sliding window of 80 bp. The averaged ratios were plotted along the genomic DNA sequence using Integrated Genome Browser (IGB) software (Affymetrix).
2011-11-01 | E-MEXP-2980 | biostudies-arrayexpress
Project description:Limb buds were dissected from E10.75 mouse embryos, fixed in 1% formaldehyde for 15 min at room temperature, washed 3 times with cold PBS and stored at -800C. Pools of 16 limb buds were used for each ChIP-chip experiment. ChIP was performed according to (Lee et al., 2006) using 2 ?g of anti-CTCF antibodies (A300-543A, Bethyl Laboratories) and EZview Red Protein G/A Affinity Gel (Sigma). Immunoprecipitated and whole cell extract DNA (input) were treated with RNaseA, proteinase K and purified by 2 rounds of extraction with phenol:chloroform: isoamyl alcohol. ChIP and input DNA were amplified using Ligation-Mediated PCR (Lee et al., 2006). PCR was limited to 15 cycles. 1.5 ?g of ChIP and input DNA were fragmented and labeled using GeneChip WT Double-Stranded DNA Terminal Labeling Kit (Affymetrix) and hybridized to the Chromosome 2 and X tiling arrays (Affymetrix). Tiling arrays data were quantile normalized within cDNA/genomic DNA or ChIP/ input replicate groups using R packages, STARR and Ringo (Zacher et al., 2010; Toedling et al., 2007). The ratio of probe intensity between the experiment and the control groups were computed considering the median values over replicates. The ratios were smoothened by computing the running medians with a half window size set to 150 bp and a minimum of 5 probes per window. To identify enriched regions a minimum of 3 consecutive probes with a smoothed ratio exceeding a threshold have been considered. The threshold has been fixed by taking the 99th percentile of the estimated null distribution of the ratios. Only ChIP enriched regions with score log2?1.5 and width ?150 bp were considered for further analysis. As a complement, array data were quantile normalized within ChIP/input replicate groups and scaled to medial feature intensity of 500 using TAS software (Affymetrix). For each genomic position, a dataset was generated consisting of all probes mapping within a sliding window of 250 bp. The averaged ratios were plotted along the genomic DNA sequence using Integrated Genome Browser (IGB) software (Affymetrix).
2011-11-01 | E-MEXP-2979 | biostudies-arrayexpress
Project description:Limb buds were dissected from E10.75 mouse embryos and stored in RNAlater reagent (Qiagen), for genotyping. For each replicate, RNA was isolated from pools of six limb buds either of wild type or homozygous mutants, using RNeasy micro-kit (Qiagen). cRNA was synthesized according to the manufacturers instructions (Ambion) and hybridized to the GeneChip Mouse Genome 430 2.0 Arrays (Affymetrix), which interrogates ca 39,000 transcripts. Three independent RNA extractions, cDNA synthesis and array hybridizations were performed. The expression arrays data were normalized and scaled to signal intensity of 100 using GCOS 1.2 software (Affymetrix). Expression levels were analyzed using GeneSpring software (Silicon Genetics, Redwood City, CA) and Matlab 2009 (Math Works, Inc., MA). To identify differentially expressed transcripts, pair-wise comparison analysis was performed with GCOS 1.2 software (Affymetrix). A 77% cutoff in consistency of change (at least seven out of nine comparisons were either increased or decreased) was applied. Only genes that satisfied the pair-wise comparison test and displayed ?1.5 fold change in expression were considered for further analysis.
2011-11-01 | E-MEXP-2977 | biostudies-arrayexpress
Project description:We wanted to identify differentially expressed genes in wild-type forelimbs and forelimbs briefly exposed to Shh signaling. E10.25 forelimbs were cultured in control media and media containing cyclopmaine (inhibits the Hedgehog pathway) Generated two wild-type (control) and two cyclopmaine biological replicates for RNA-seq UT-Genome and Analysis Facility
2015-07-01 | E-GEOD-58664 | biostudies-arrayexpress
Project description:Spatiotemporal regulation of GLI target genes in the mammalian limb bud
| PRJNA253126 | ENA
Project description:During limb development, Hoxd genes are transcribed in two waves: Early on, when the arm and forearm are specified and subsequently, when digits form. While the latter phase is controlled by enhancers centromeric to the HoxD cluster, we show here that the early phase requires enhancers located in the opposite telomeric gene desert. The transition between the two types of regulations involves a functional switch between two distinct topological domains, as reflected by a subset of genes mapping centrally into the cluster, which initially interact with the telomeric domain and subsequently shift to establish new contacts on the opposite side. This transition between two regulatory landscapes generates an intermediate area of low Hox dose developing into the wrist, the transition between our arms and our hands. This intriguing correspondence between genomic and morphological boundaries illustrates the mechanism underlying collinear Hox gene regulation in our developing appendages. Chromatin ImmoPrecipitation on chip (Tiling array): Distribution of H3K4me3 and H3K27me3 in early limb buds at E9.5, E10.5 and proximal late limbs E12.5. Distribution of H3K27me3 in del(8-13) and del(8-13)/del(attP-TpSB3) E10.5 limb buds. Distribution of H3K27me3 in WT and homozygote del(Nsi-Atf2) (Montavon et al., 2011) forelimb autopods.
2013-06-07 | E-GEOD-45454 | biostudies-arrayexpress
Project description:During limb development, Hoxd genes are transcribed in two waves: Early on, when the arm and forearm are specified and subsequently, when digits form. While the latter phase is controlled by enhancers centromeric to the HoxD cluster, we show here that the early phase requires enhancers located in the opposite telomeric gene desert. The transition between the two types of regulations involves a functional switch between two distinct topological domains, as reflected by a subset of genes mapping centrally into the cluster, which initially interact with the telomeric domain and subsequently shift to establish new contacts on the opposite side. This transition between two regulatory landscapes generates an intermediate area of low Hox dose developing into the wrist, the transition between our arms and our hands. This intriguing correspondence between genomic and morphological boundaries illustrates the mechanism underlying collinear Hox gene regulation in our developing appendages. Chromatin ImmunoPrecipitation and Sequencing (ChIP-seq) of H3K27A in developing proximal and distal limbs at E9.5, E10.5 and E12.5
2013-06-07 | E-GEOD-45456 | biostudies-arrayexpress