Project description:We have adapted ATAC-seq to sea urchin embryos. As a proof of concept, we documented strong ATAC-seq signals over the locations of a number of previously authenticated, developmentally active, cell type specific cis-regulatory modules. We then used the method to generate a systematic developmental series for embryos from three independent batches, obtained every six hours from blastula stage until completion of embryogenesis at 72h. The independent batches provide internal reproducibility controls.
Project description:An increasing number of studies, including mutant expression profiling and comparative transcriptomic analyses, require reference RNA-seq data collections in mice. Particularly, to complement previous profiling data sets based on arrays, a full RNA-seq developmental series will be required for whole embryos. E10.5 is a key reference stage as it represents the early organogenesis stage. Here, we have performed high-throughput sequencing of total RNA form whole mice embryos at embryonic stage E10.5.
Project description:An increasing number of studies, including mutant expression profiling and comparative transcriptomic analyses, require reference RNA-seq data collections in mice. Particularly, to complement previous profiling data sets based on arrays, a full RNA-seq developmental series will be required for whole embryos. E10.5 is a key reference stage as it represents the early organogenesis stage. Here, we have performed high-throughput sequencing of total RNA form whole mice embryos at embryonic stage E10.5. Sequencing of the total RNA of whole embryos of mouse at embryonic stage E10.5.
Project description:Purpose: The Tbrain transcription factor has demonstrated an evolved preference for low-affinity, secondary site binding motifs between the sea star and sea urchin orthologs. We sought to identify targets of sea urchin and sea star orthologs of Tbr. Because less is known about the function of Tbr during sea star development, we used RNA-seq in conjuction with ChIP-seq studies (GEO:xxxx) to determine the targets of sea star Tbr in early development. Methods: Sea star (Patiria miniata) embryos were injected with translation-blocking morpholino antisense oligonucleotides to knock-down PmTbr expression, as described previously. Control morpholinos were injected into sibling embryos. Embryos were allowed to develop until hatching (30-36 hpf) at which point injected embryos were collected and RNA was extracted. RNA-seq libraries were prepared, sequenced, and analyzed using standard protocols. Results: There are 2,562 genes that are significantly differentially expressed relative to control morpholino inected embryos (FDR < 0.05). There are roughly equivalent numbers of genes down-regulated (1,041) and up-regulated (1,521) by Pm-tbr knockdown, suggesting that PmTbr may act as both a transcriptional activator and repressor. 1,165 differentially expressed genes are located within 75 kb of a PmTbr binding site determined using ChIP-seq, and this set is used as a basis for comparison between sea star and sea urchin binding sites. Conclusions: 1,165 targets of the PmTbr transcription factor were identified based on differential expression following knockdown and the presence of transcription factor binding sites proximal to differentially expressed genes. There are an equal number of up- and down-regulated targets, suggesting Tbr may function as a transcriptional activator and repressor, depending on context and target gene. There was no clear association of motif utilization with either the direction of differential expression or ontological category of the target gene. There are only a small fraction of target genes (approximately 10%) that are in common between the sea star and sea urchin sets.
Project description:Transcription factor SoxC is required for all neural development in purple sea urchin S. purpuratus embryos. To begin to develop a gene regulatory network for neural development, we used RNA-Seq to compare transcript populations in SoxC knockdown and control embryos.
Project description:Pre-implantation embryogenesis encompasses several critical events including genome reprogramming, zygotic genome activation (ZGA), and cell fate commitment, of which most remain mechanistically unclear in primate. Here we carried out time-series RNA-seq in 26 single and 8 pooled rhesus monkey oocytes and pre-implantation embryos encompassing representative developmental stages to explore these process.
Project description:The transcriptome of E9.5 embryos lacking functional RNase H2 (Rnaseh2bE202X/E202X) was compared to age-matched wild type controls to investigate the molecular basis of growth arrest in Rnaseh2-/- mice. Total RNA extracted from three E9.5 embryos of Rnaseh2+/+ and Rnaseh2E202X/E202X were compared
Project description:Animal embryos have the remarkable property of self-organization. Over 125 years ago Hans Driesch separated the two blastomeres of sea urchin embryos and obtained twins, in what was the foundational experiment of experimental embryology. Since then, embryonic twinning has been obtained experimentally in many animals by diverse methods. In a recent study, we developed bisection methods that generate identical twins reliably from Xenopus blastula embryos. In the present study we investigated the transcriptome of regenerating half-embryos after sagittal and dorsal-ventral (D-V) bisections. Individual embryos were operated at midblastula with an eyelash hair and cultured until early gastrula (stage 10.5) or late gastrula (Stage 12) and analyzed the transcriptome of each half-embryo by RNAseq. Because many genes are activated by wound healing, stringent analyses were used to identify genes upregulated in identical twins but not in either dorsal or ventral fragments. At early gastrula cell division-related genes such as histones were identified, whereas at late gastrula pluripotency genes (such as sox2) and germ layer determining genes (such as eomesodermin, ripply2 and activing receptor ACVRI) and a number of secretory pathway components (serpinH1, fucoleptin and sialyl transferase). These findings are consistent with a model in which cell division is required to heal damage, while maintaining pluripotency to permit formation of the organizer with a displacement of 900 from its original site. In addition, the extensive transcriptomic data presented here (30 RNA-seq libraries of individual whole or regenerating half-embryos) provides a useful resource for data mining gene expression during early vertebrate development.
Project description:Embryonic stem (ES) cells and embryos reversibly pause via chemical mTOR inhibition. In this study, we investigate the tissue-specific response to mTORi-induced pausing in ES and trophoblast stem (TS) cells. To resolve the sequential rewiring of the proteome, we conducted a time-series proteomics experiment at 1, 3, 6, 12, 24, and 48 hours upon induction of pausing, and at 1, 3, 6, 12, 24, and 48 hours upon release of pausing in ES and TS cells. We find that ES, but not TS cells pause reversibly. To optimise developmental pausing conditions, we reasoned that by understanding the difference in pausing response of ES and TS cells, we could identify which pathways are essential for pausing. We found that KEGG pathways related to amino acid degradation, fatty acid degradation, and DNA repair are upregulated in ES cells, but downregulated in TS cells during entry into pausing. Moreover, by targeted metabolomics, we found a depletion of short chain carnitines in the paused ES cells. To extend the length of developmental pausing, we supplemented paused embryos with L-carnitine. The L-carnitine supplementation facilitates lipid usage and prolongs the pausing length by 19 days through the establishment of a more dormant state.
Project description:The transcriptome of E9.5 embryos lacking functional RNase H2 (Rnaseh2bE202X/E202X) was compared to age-matched wild type controls to investigate the molecular basis of growth arrest in Rnaseh2-/- mice.