Transcriptome profiling of cytosol, endoplasmic reticulum (ER), and pellet fraction from the Xenopus laevis oocyte and mature egg
Ontology highlight
ABSTRACT:
ORGANISM(S): Xenopus laevis
PROVIDER: GSE199254 | GEO | 2022/12/13
REPOSITORIES: GEO
Similar Datasets
Project description:As an important post-transcriptional regulatory mechanism, asymmetric localization of mRNAs is essential for cell polarity and cell fate determination during early development. In an effort to understand localization of mRNAs encoding proteasome components during Xenopus oocyte-to-embryo transition, we discovered that the endoplasmic reticulum (ER), which is thought to mainly serve “house-keeping” roles in the cell, plays a multifaceted role in controlling localization of maternal RNAs. Our RNA-seq analysis of fractionated transcripts reveals that more than 40% of RNAs, including proteasome mRNAs, are present on the ER in Xenopus oocytes. After meiotic maturation, a large fraction of ER-associated RNA is released into the cytosol. The release of proteasome RNAs from the ER into the cytosol is conserved during mouse oocyte maturation. Our comparative proteomic analysis and ribonucleoprotein immunoprecipitation demonstrate that the majority of ER-associated RNA-binding proteins (RBPs) remain associated with the ER after oocyte maturation. However, all ER-associated RBPs analyzed exhibit reduced binding to some of their target RNAs after oocyte maturation, providing a mechanistic explanation for the dynamic regulation of RNA-ER association. We further show that the ER is remodeled massively during Xenopus oocyte maturation, leading to the formation of a widespread tubular ER network in the animal hemisphere that is required for the asymmetric localization of proteasome mRNAs in mature eggs. To our knowledge, our findings demonstrate for the first time that dynamic regulation of RNA-ER association and remodeling of the ER are fundamentally important for asymmetric localization of RNAs during development.
2022-12-15 | PXD033018 | Pride
Project description:Mass spectrometry based proteomics enables the global identification and quantification of proteins and their post-translational modifications in complex biological samples. Proteomic analysis requires a complete and accurate reference set of proteins, and is therefore largely restricted to model organisms with sequenced genomes. Here, we demonstrate the feasibility of deep genome-free proteomics using a reference proteome derived from heterogeneous mRNA data. We identify more than 11k proteins with 99% confidence from the unfertilized, X. laevis egg and estimate protein abundance with approximately two-fold precision. Our reference database outperforms the provisional gene models based on genomic DNA-sequencing and references generated by other methods. Surprisingly, we find that many proteins in the egg lack mRNA support and many of these proteins are found in blood or liver, suggesting that they are taken up from the blood plasma, together with yolk, during oocyte growth and maturation, potentially contributing to early embryogenesis. To facilitate proteomics in non-model organisms, we make our platform available as an online resource which converts heterogeneous mRNA data into a protein reference set. Thus, we demonstrate the feasibility and power of genome-free proteomics while shedding new light on embryogenesis in vertebrates.
2014-05-30 | PXD000926 | Pride
Project description:Wood-Pellet and Cork Biofilters
| PRJNA821383 | ENA
Project description:identification of proteins in polysome pellet with and without silvestrol treatment
2014-09-16 | PXD001310 | Pride
Project description:The genetic causes of oocyte meiotic deficiency (OMD), a form of primary infertility characterised by the production of immature oocytes, remain largely unexplored. Using whole exome sequencing, we found that 26% of a cohort of 23 subjects with OMD harboured the same homozygous nonsense pathogenic mutation in PATL2, a gene encoding a putative RNA-binding protein. Using Patl2 knockout mice, we confirmed that PATL2 deficiency disturbs oocyte maturation, since oocytes and zygotes exhibit morphological and developmental defects respectively. PATL2's amphibian orthologue is involved in the regulation of oocyte mRNA as a partner of CPEB. However, Patl2's expression profile throughout oocyte development in mice, alongside colocalisation experiments with Cpeb1, Msy2 and Ddx6 (three oocyte RNA-regulators) suggest an original role for Patl2 in Mammals. Accordingly, transcriptomic analysis of oocytes from WT and Patl2-/- animals demonstrated that in the absence of Patl2, expression levels of a select number of highly relevant genes involved in oocyte maturation and early embryonic development are deregulated. In conclusion, PATL2 is a novel actor of mammalian oocyte maturation whose invalidation causes OMD in humans.
| EGAS00001002903 | EGA
Project description:In eukaryotic cells, the spatial regulation of protein expression is frequently conferred through the coupling of mRNA localization and the local control of translation. mRNA localization to the endoplasmic reticulum (ER) is a prominent example of such regulation and serves a ubiquitous role in segregating the synthesis of secretory and integral membrane proteins to the ER. Recent genomic and biochemical studies have now expanded this view to suggest a role for the ER in global protein synthesis. We have utilized cell fractionation and ribosome profiling to obtain a genomic survey of the subcellular organization of mRNA translation and report that ribosomal loading of mRNAs, a proxy for mRNA translation, is biased to the ER. Notably, ER-associated mRNAs encoding both cytosolic and topogenic signal-encoding proteins display similar ribosome loading densities, suggesting that ER-associated ribosomes serve a global role in mRNA translation. We propose that the partitioning of mRNAs and their translation between the cytosol and ER compartments may represent a novel mechanism for the post-transcriptional regulation of gene expression.
2011-12-20 | GSE31539 | GEO
Project description:In several metazoans the number of active replication origins in embryonic nuclei is higher than in somatic ones, ensuring rapid genome duplication during synchronous embryonic cell divisions. High replication origin density can be restored by somatic nuclear reprogramming. However, mechanisms underlying high replication origin density formation coupled to rapid cell cycles are poorly understood. Here, we characterized a fraction of Xenopus egg extract able to stimulate somatic DNA replication.
2020-03-19 | PXD017383 | Pride
Project description:Oocyte maturation, fertilization, and early embryonic development occur in the absence of gene transcription. Therefore, it is critical to understand at a global level the post-transcriptional events that are driving these transitions. Here, we have used a systems approach by combining polysome mRNA profiling and bioinformatics to identify RNA binding motifs in mRNAs that either enter or exit the polysome pool during mouse oocyte maturation. Association of mRNA with the polysomes correlates with active translation.
2012-01-14 | GSE35106 | GEO
Project description:In eukaryotic cells, the spatial regulation of protein expression is frequently conferred through the coupling of mRNA localization and the local control of translation. mRNA localization to the endoplasmic reticulum (ER) is a prominent example of such regulation and serves a ubiquitous role in segregating the synthesis of secretory and integral membrane proteins to the ER. Recent genomic and biochemical studies have now expanded this view to suggest a role for the ER in global protein synthesis. We have utilized cell fractionation and ribosome profiling to obtain a genomic survey of the subcellular organization of mRNA translation and report that ribosomal loading of mRNAs, a proxy for mRNA translation, is biased to the ER. Notably, ER-associated mRNAs encoding both cytosolic and topogenic signal-encoding proteins display similar ribosome loading densities, suggesting that ER-associated ribosomes serve a global role in mRNA translation. We propose that the partitioning of mRNAs and their translation between the cytosol and ER compartments may represent a novel mechanism for the post-transcriptional regulation of gene expression. HEK293 cells were fractionated between the cytosol and endoplasmic reticulum. Within each fraction, ribosome footprints were generated and sequenced. In parallel, total mRNA was sequenced.
2011-12-20 | E-GEOD-31539 | biostudies-arrayexpress
Project description:Xenopus egg extract represents a powerful cell-free biochemical tool for studying organelle assembly and function. Large quantities of cytoplasm can be isolated, and biochemical manipulation of extract composition and cell cycle state is relatively straightforward. In this protocol, we describe the reconstitution of nuclear assembly by adding a chromatin source to interphasic X. laevis egg extract. Intact nuclei assemble within 30-45 min of initiating the reaction, followed by nuclear growth. We also describe methods for imaging and quantifying nuclear import kinetics. Recombinant proteins or small molecules of interest can be added to the extract before or after nuclear assembly, and immunodepletion allows for removal of specific proteins from the extract. This approach will continue to inform mechanisms of nuclear assembly, nuclear pore complex assembly and function, nucleocytoplasmic transport, DNA replication, nuclear envelope breakdown, and nuclear size and shape regulation.
| S-EPMC6540748 | biostudies-literature