A mesodermal gene regulatory network directed by zebrafish No tail
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE12331 | GEO | 2009/02/19
SECONDARY ACCESSION(S): PRJNA113221
REPOSITORIES: GEO
Similar Datasets
Project description:Using chromatin immunoprecipitation combined with microarrays we have identified targets of No tail (Ntl), a zebrafish Brachyury ortholog that plays a central role in mesoderm formation. We show that Ntl regulates a downstream network of other transcription factors and identify an in vivo Ntl binding site that resembles the consensus T-box binding site (TBS) previously identified by in vitro studies. We show floating head (flh) is a direct transcriptional target of Ntl in the notochord and that TBS in the flh upstream region are required for Ntl directed expression. Using this genome-scale data we have assembled a Gene Regulatory Network that describes mesoderm formation and patterning in the early zebrafish embryo and which predicts a role for Ntl in somite segmentation. Keywords: ChIP-chip Genomic array design. Microarrays were designed as described below and manufactured by Agilent Technologies (www.agilent.com). We have previously described the ChIP-chip technique in zebrafish and the design of promoter microarrays which cover a 2kb region around the transcription start sites (TSSs) of 11,512 zebrafish genes based on Zv4 (Wardle et al, 2006. Genome Biology 7:R71). In order to capture additional regulatory information further from the basal promoter region we also designed an expanded set of 60mer probes that cover 9kb upstream and 3kb downstream of the TSS of these genes using the same parameters as for the 2kb microarrays. We also incorporated several sets of control probes, both positive and negative. On each array there are 753 negative probes designed against zebrafish gene desert regions and Arabidopsis thaliana genes. In addition we included duplicates of promoter probes for several genes that we anticipated may be bound by factors of interest for this and other studies (wnt11, flh, vent, msgn1, myod, fgf8, pcdh8) and these probes are arrayed on both slides slide. We also included in this design 502 intensity control probes. Finally there are 2,256 controls added by Agilent. The final design contained 378,002 probes, including 365,537 experimental probes divided between nine microarray slides: slides 1-8 contained 40,616 experimental probes and slide 9 contained 40, 609. Further information on design and manufacture of the microarrays can be found on the Agilent Technologies website (www.agilent.com).
2009-02-19 | E-GEOD-12331 | biostudies-arrayexpress
Project description:The zebrafish genes spadetail (spt) and no tail (ntl) encode T-box transcription factors that are important for early mesoderm development. Although much has been done to characterize these genes, the identity and location of target regulatory elements remain largely unknown. Here, we survey the genome for downstream target genes of the Spt and Ntl T-box transcription factors. We find evidence for extensive additive interactions towards gene activation and limited evidence for combinatorial and antagonistic interactions between the two factors. Using in vitro binding selection assays to define Spt- and Ntl-binding motifs, we searched for target regulatory sequence via a combination of binding motif searches and comparative genomics. We identified regulatory elements for tbx6 and deltaD, and, using chromatin immunoprecipitation, in vitro DNA binding assays and transgenic methods, we provide evidence that both are directly regulated by T-box transcription factors. We also find that deltaD is directly activated by T-box factors in the tail bud, where it has been implicated in starting the segmentation clock, suggesting that spt and ntl act upstream of this process. Keywords: Morpholino Knockdown We measured transcript levels in zebrafish embryos at 75% epiboly. We compared levels in spadetail (spt) and/or no tail (ntl) morpholino injected embryos to those in control embryos mock injected with Danieau solution plus 0.25% phenol red. Three biological replicates are included for each treatment.
2010-05-19 | E-GEOD-12857 | biostudies-arrayexpress
Project description:We screened for differentially expressed genes in the developing notochord using the Affymetrix microarray system in Xenopus laevis. At late gastrula, we dissected four regions from the embryo, anterior mesoderm, posterior mesoderm, notochord and presomitic mesoderm. Three types of comparison were carried out to generate a list of predominantly notochord expressed genes: (1) Posterior mesoderm vs. anterior mesoderm; notochord genes are expected to be increased since the notochord is located in the posterior mesoderm. (2) Posterior mesoderm vs. whole embryos; notochord genes are expected to be increased. (3) Notochord vs. somite. This comparison sub-divided the group of posterior mesodermal genes identified in (1) and (2). All tissues are dissected using tungsten needles. We first dissected dorsal tissue above the archenteron from late gastrula to early neurula. To loosen tissue, we treated the dissected dorsal explant in a 1% cysteine solution (pH 7.4) and removed the neuroectodermal layer. Anterior mesoderm was dissected corresponding to about the anterior one-third of the archenteron roof, and the rest was collected as posterior mesoderm. The posterior mesodermal explant was dissected into notochord and somites, following a clearly visible border between the two tissues. The accuracy of all dissection was confirmed by RT-PCR of marker genes.
2006-12-31 | E-MEXP-717 | biostudies-arrayexpress
Project description:The zebrafish genes spadetail (spt) and no tail (ntl) encode T-box transcription factors that are important for early mesoderm development. Although much has been done to characterize these genes, the identity and location of target regulatory elements remain largely unknown. Here, we survey the genome for downstream target genes of the Spt and Ntl T-box transcription factors. We find evidence for extensive additive interactions towards gene activation and limited evidence for combinatorial and antagonistic interactions between the two factors. Using in vitro binding selection assays to define Spt- and Ntl-binding motifs, we searched for target regulatory sequence via a combination of binding motif searches and comparative genomics. We identified regulatory elements for tbx6 and deltaD, and, using chromatin immunoprecipitation, in vitro DNA binding assays and transgenic methods, we provide evidence that both are directly regulated by T-box transcription factors. We also find that deltaD is directly activated by T-box factors in the tail bud, where it has been implicated in starting the segmentation clock, suggesting that spt and ntl act upstream of this process. Keywords: Morpholino Knockdown
2008-09-20 | GSE12857 | GEO
Project description:Here, using ChIP-Seq, we examined the targets of Nanog-like and Mxtx2 in blastula stage zebrafish embryos. We found that Nanog-like bind to its known targets like Oct4, Sox2, and Nanog-like. Nanog-like also bound to genes involved in extraembryonic lineage differentiation, like gata3 and krt4 for EVL differentiation, and mxtx2 and slc26a1 for YSL differentiation, mesoderm specification like ntl and tbx3, cell movement like wnt11 and cxcr4b, and signaling genes like ndr1, bmp2b, fgf8a and wnt8a. The binding profile suggests that Nanog-like may play a versatile role involving many developmental processes. We found 11.3% of the genes (1751 out of all annotated 15500 zebrafish genes) and 43.6% of the YSL genes (118 out of 271 genes expressed in the YSL) were bound by Mxtx2, suggesting Mxtx2 bound directly to YSL genes to activate their expression Examination of Nanog-like and Mxtx2 binding sites in 3.5hpf and 4.5hpf zebrafish embryos
2011-12-28 | E-GEOD-34683 | biostudies-arrayexpress
Project description:Oncogenic transformation of individual cell fates by developmental signaling cascades and transcription factors triggers diverse cancer types. Chordoma is a rare, aggressive tumor arising from transformed notochord remnants. Various potentially oncogenic factors have been found deregulated in chordoma and its metastases, yet clear causation remains uncertain. In particular, expression of the notochord-controlling transcription factor Brachyury is hypothesized as key molecular driver in chordoma formation, yet an in vivo model to causally test its oncogenic potential in the notochord is missing. Here, we apply a zebrafish model of chordoma onset to identify the notochord-transforming potential of tumor-implicated candidate genes in vivo. We find that overexpression of human and zebrafish Brachyury, including a version with augmented transcriptional activity, is insufficient to initiate notochord hyperplasia in vivo. In contrast, the repeatedly chordoma-implicated receptor tyrosine kinase (RTK) genes EGFR and KDR/VEGFR2 are sufficient to transform developmental notochord cells, akin to direct activation of Ras. Analysis of transcriptome and sub-cellular organization from transformed notochords suggests that aberrant activation of RTK/Ras signaling attenuates processes required for the differentiation of notochord cells. Taken together, our results provide first in vivo indication for a lack of tumor-initiating potential of Brachyury expression in the notochord, and suggest activated RTK signaling as potent hyperplasia-initiating event in chordoma.
2018-12-01 | E-MTAB-7349 | biostudies-arrayexpress
Project description:Here, using ChIP-Seq, we examined the targets of Nanog-like and Mxtx2 in blastula stage zebrafish embryos. We found that Nanog-like bind to its known targets like Oct4, Sox2, and Nanog-like. Nanog-like also bound to genes involved in extraembryonic lineage differentiation, like gata3 and krt4 for EVL differentiation, and mxtx2 and slc26a1 for YSL differentiation, mesoderm specification like ntl and tbx3, cell movement like wnt11 and cxcr4b, and signaling genes like ndr1, bmp2b, fgf8a and wnt8a. The binding profile suggests that Nanog-like may play a versatile role involving many developmental processes. We found 11.3% of the genes (1751 out of all annotated 15500 zebrafish genes) and 43.6% of the YSL genes (118 out of 271 genes expressed in the YSL) were bound by Mxtx2, suggesting Mxtx2 bound directly to YSL genes to activate their expression
2011-12-28 | GSE34683 | GEO
Project description:Xenopus embryonic ectodermal cells are responsive to various inducing factors. Mesoderm is specified and patterned by extracellular factors including FGF, Nodal, BMP and Wnt families. Pinhead is another secreted protein implicated in mesoderm formation. We found that Pinhead physically interacts with and antagonizes ADMP (anti-dorsalizing morphogenetic protein) that acts as BMP-like protein to promote ventral mesoderm formation. ADMP and BMPs have been shown to cooperate to activate phospho-Smad1 signaling to lead to ventral mesoderm development. Since Chordin is a BMP antagonist, we hypothesized that Pinhead together with Chordin can promote downregulation of phospho-Smad1 signaling that leads to dorsal mesoderm development. RNA sequencing revealed that Pinhead and Chordin synergize in dorsal mesoderm formation in ectodermal explants.
2021-04-19 | GSE168370 | GEO
Project description:Transcription factors play diverse roles during embryonic development, combinatorially controlling multiple cellular states in a spatially and temporally defined manner. Resolving the dynamic transcriptional profiles that underlie these patterning processes is essential for understanding embryogenesis at the molecular level. Here we show how temporal, tissue-specific changes in embryonic transcription factor function can be discerned by integrating caged morpholinos (cMOs) with photoactivatable fluorophores, fluorescence-activated cell sorting (FACS), and microarray technologies. As a proof of principle, we have dynamically profiled No tail-a (Ntla)-dependent genes at different stages of axial mesoderm development in zebrafish, discovering and characterizing discrete sets of transcripts that are coincident with either notochord cell fate commitment or differentiation. Our studies demonstrate how optically controlled chemical tools can be use to probe developmental processes with spatiotemporal precision and reveal the sequential activation of distinct transcriptomes within a cell lineage by a single transcriptional factor. Zebrafish zygotes were injected with a mixture of ntla caged morpholino (cMO) and caged fluorescein dextran (cFD), and a 100 µm-diameter region of the posterior chordamesoderm 125 μm anterior to the center of Kupffer’s vesicle was UV-irradiated at 12 hours post fertilization (hpf). The UV irradiation generated an active morpholino targeting the ntla 5'UTR and simultaneously labeled the cells with green fluorescence. By 36 hpf, the irradiated, green-fluorescent cells contributed to both notochord and floor plate, but the notochord cells were partially vacuolated and disorganized. Combining these caged reagents with microarray analysis identified transcriptional changes coincident with loss of ntla at 16 hpf and subsequent notochord differentiation.
2012-03-01 | GSE31881 | GEO
Project description:Purpose: To construct cell landscape of graded Nodal signaling in zebrafish explants. Methods: Nodal injected explants (injected with 10pg ndr2 mRNA) were harvested at 4hpf, 5hpf, 6hpf, 8hpf, 10hpf, 24hpf. Libraries were prepared using Chromium Controller and Chromium Single Cell 3’Library & Gel Bead Kit v2 (10x Genomics, PN-120237) according to the manufacturer’s protocol for 10000 cells recovery. Results: A total of 42,317 single-cell transcriptomes were collected after stringent quality control measures. Conclusions: 19 Cell types from 6 developmental stages were identified, Nodal explant mainly contains the axial mesoderm (prechordal plate and notochord) and anterior-posterior patterned head structures.
2021-01-28 | GSE165653 | GEO