Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

A Genome-Wide Survey of Maternal and Embryonic Transcripts during Xenopus tropicalis Development

ABSTRACT: To analyze the dynamics and diversity of coding and non-coding transcripts during development, both polyadenylated mRNA and ribosomal RNA-depleted total RNA were harvested across six developmental stages and subjected to high throughput sequencing. The maternally loaded transcriptome is highly diverse and consists of both polyadenylated and deadenylated transcripts. Many maternal genes show peak expression in the oocyte and include genes which are known to be the key regulators of events like oocyte maturation and fertilization. Of all the transcripts that increase in abundance between an early blastula and larval stages, about 30% of the embryonic genes are induced by fourfold or more by the late blastula stage and another 35% by late gastrulation. Using a gene model validation and discovery pipeline, we identified novel transcripts, the majority of which show hallmarks of being long non-coding RNAs both in terms of their coding potential and gene structure. Profiles of polyadenylated mRNA (6 stages) and ribosomal RNA-depleted total RNA (3 stages) through early Xenopus tropicalis development

ORGANISM(S): Xenopus (Silurana) tropicalis

SUBMITTER: Gert Jan Veenstra

PROVIDER: E-GEOD-43652 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

A genome-wide survey of maternal and embryonic transcripts during Xenopus tropicalis development.

Paranjpe Sarita S SS Jacobi Ulrike G UG van Heeringen Simon J SJ Veenstra Gert Jan C GJ

BMC genomics 20131106

<h4>Background</h4>Dynamics of polyadenylation vs. deadenylation determine the fate of several developmentally regulated genes. Decay of a subset of maternal mRNAs and new transcription define the maternal-to-zygotic transition, but the full complement of polyadenylated and deadenylated coding and non-coding transcripts has not yet been assessed in Xenopus embryos.<h4>Results</h4>To analyze the dynamics and diversity of coding and non-coding transcripts during development, both polyadenylated mR ...[more]

PMID: 24195446

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Project description:The transformation of a fertilized oocyte into a multicellular embryo comprised of differentiated cells depends on the processing of genetic orders, first in the form of transcripts and then of proteins. Our current understanding of mouse development has greatly benefited from the analysis of transcripts as proxy for the proteins. However, the hexagonal-shaped free ribosomes that translate mRNAs into proteins are scarce during cleavage and do not become abundant until the morula-blastocyst stage. How accurate, then, is an understanding of mouse and more in general mammalian development that uses the analysis of transcripts as proxy for the proteins? This led us in this study to measure the proteome of seven preimplantation mouse stages (B6C3F1 x CD1), from oocyte to blastocyst, using 'spike-in' SILAC technology combined with high accuracy mass spectrometry (LTQ Orbitrap and Q-Exactive). The relative abundances of embryonic proteins were compared and contrasted with those of transcripts measured by RNA sequencing. A total of 6976 proteins were detected in at least the spike (precondition for determining the heavy/light peptide ratios in oocytes or embryos). Among these 6976 proteins, 4991 proteins were present in all developmental stages, and 1893 proteins were present in all replicates. Our tandem approach uncovered: 1) the different abundance profiles of proteome and transcriptome during the time of preimplantation development; 2) the different reciprocal associations of developmental stages (dendrograms) on the protein level as compared to the mRNA level; and 3) the different gene ontology terms overrepresented among the proteins that increase or decrease across adjacent stages, compared to mRNAs. In essence, the elaborate phasing of embryonic gene expression that has been known for mRNAs leaves an unsuspected footprint on the protein products of those mRNAs. This information facilitates the analysis of mammalian development in two important ways. First, it provides new insight into the regulation of the transition from the differentiated oocyte into the embryo, by identifying the subsets of proteins that accompany the passage from one embryonic stage to the next, which are likely composed of important regulators of the morphogenetic transitions. Second, our analysis provides a reference to determine the degree of ‘embryoness’ of entities produced via assisted reproductive technologies, cloning by somatic cell nuclear transfer, and even artificial gametes.

Project description:During development histone modifying enzymes are required for cell identity and lineage commitment, however little is known about the regulatory origins of the epigenome during embryonic development. Here we generate a comprehensive set of embryonic epigenome reference maps, which we use to determine the extent to which maternal factors shape chromatin state in Xenopus embryos. Using Î±-amanitin to inhibit zygotic transcription, we find that the majority of H3K4me3 and H3K27me3-enriched regions form a maternally defined epigenetic regulatory space with an underlying logic of hypomethylated islands. This maternal regulatory space extends to a substantial proportion of neurula stage-activated promoters. In contrast, p300-recruitment to distal regulatory regions requires embryonic transcription at most loci. The results show that H3K4me3 and H3K27me3 are part of a regulatory space that exerts an extended maternal control well into post-gastrulation development, and highlight the combinatorial action of maternal and zygotic factors through proximal and distal regulatory sequences. We have performed ChIP-sequencing of eight histone modifications, RNA polymerase II (RNAPII) and the enhancer protein p300 at five stages of development: blastula (st. 9), gastrula (st. 10.5, 12.5), neurula (st. 16) and tailbud (st. 30). These experiments allow identification of enhancers (H3K4me1, p300), promoters (H3K4me3, H3K9ac), transcribed regions (H3K36me3, RNAPII) and repressed and heterochromatic domains (H3K27me3, H3K9me2, H3K9me3, H4K20me3). In addition we generated pre-MBT (st. 8) maps for three histone modifications (H3K4me3, H3K9ac, H3K27me3) and single-base resolution DNA methylome maps using whole genome bisulfite sequencing of blastula and gastrula (st. 9 and 10.5) embryos. To determine the maternal and zygotic contributions to chromatin state, we used alpha-amanitin to block embryonic transcription. Fertilised eggs were injected with 2.3 nl of 2.67 ng/ul alpha-amanitin and developed until the control embryos reached mid-gastrulation. Alpha-amanitin and control embryos were used for RNA-seq and ChIP-seq of RNAPII, H3K4me3, H3K27me3 and p300. For all ChIP-seq samples of the epigenome reference maps and RNAPII ChIP-seq samples of the Î±-amanitin experiments three biological replicates of different chromatin isolations of 45 embryos were pooled. Two biological replicates for H3K4me3 (Î±-amanitin injected: resp. 90 and 56 embryo equivalents (eeq); control: resp. 45 and 67 eeq), H3K27me3 (Î±-amanitin injected: resp. 90 and 180 eeq; control: resp. 45 and 202 eeq) and p300 (Î±-amanitin injected: resp. 112 and 56 eeq; control: resp. 112 and 67 eeq) ChIP-seq samples of the Î±-amanitin experiments were generated. For RNA-seq samples of the Î±-amanitin experiments RNA from 5 embryos from one biological replicate was isolated and depleted of ribosomal RNA

Dataset Information

A Genome-Wide Survey of Maternal and Embryonic Transcripts during Xenopus tropicalis Development

Publications

A genome-wide survey of maternal and embryonic transcripts during Xenopus tropicalis development.

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