Dataset Information

DIS3L, cytoplasmic exosome catalytic subunit, is essential for development but not cell viability in mice.

ABSTRACT: Among numerous enzymes involved in RNA decay, processive exoribonucleases are the most prominent group responsible for the degradation of the entire RNA molecules. The role of mammalian cytoplasmic 3’-5’ exonuclease DIS3L at the organismal level remained unknown. Herein we established knock-in and knock-out mouse models to study DIS3l functions in mice. DIS3L is indeed a subunit of the cytoplasmic exosome complex, which disruption leads to severe embryo degeneration and death in mice soon after implantation. These changes could not be prevented by supplementing extraembryonic tissue with functional DIS3L through the construction of chimeric embryos. Preimplantation DIS3L KO embryos were unaffected in their morphology and ability to produce functional embryonic stem cells (ES) cells showing that DIS3L is not essential for cell viability. There were also no major changes in the transcriptome level for both ES cells and blastocysts, as revealed by RNA sequencing experiments. Notably, however, DIS3l KO led to inhibition of the global protein synthesis. These results point to the essential role of DIS3l in mRNA quality control pathways crucial for proper protein synthesis during embryo development.

INSTRUMENT(S):

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Dominik Cysewski

LAB HEAD: Andrzej Dziembowski

PROVIDER: PXD038745 | Pride | 2025-05-06

REPOSITORIES: Pride

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			Action	DRS
	80309513mro_B-cells-WT1.raw	Raw
	80309514mro_B-cells-46CGFP1.raw	Raw
	80309515mro_B-cells-WT2.raw	Raw
	80309516mro_B-cells-46CGFP2.raw	Raw
	80309517mro_BMDM-WT.raw	Raw

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DIS3L, cytoplasmic exosome catalytic subunit, is essential for development but not cell viability in mice.

Brouze Michał M Szpila Marcin M Czerwińska Areta A Antczak Wiktor W Mroczek Seweryn S Kuliński Tomasz M TM Hojka-Osińska Anna A Cysewski Dominik D Gewartowska Olga O Adamska Dorota D Gruchota Jakub J Borsuk Ewa E Dziembowski Andrzej A

RNA (New York, N.Y.) 20250416 5

Among numerous enzymes involved in RNA decay, processive exoribonucleases are the most prominent group responsible for the degradation of entire RNA molecules. The role of mammalian cytoplasmic 3'-5' exonuclease DIS3L at the organismal level remained unknown. Herein, we established knock-in and knockout (KO) mouse models to study DIS3L functions in mice. DIS3L in mice is indeed a subunit of the cytoplasmic exosome complex, the disruption of which leads to severe embryo degeneration and death in ...[more]

PMID: 39919786

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Project description:Embryonic stem cells (ES cells) are pluripotent stem cells that can contribute to all lineages of the embryo proper and were first isolated in 1981 in this laboratory (Evans and Kaufmann, 1981). They have contributed immeasurably to biology, both in terms of studying pluripotent stem cell maintenance in culture and in vitro differentiation, and because they can be easily genetically manipulated in culture, allowing for the production of mice with designed specific mutations (targeted mice), via homologous recombination. The recent isolation of human ES cell lines is promising in terms of stem cell therapy development. Mouse ES cell lines are isolated from the inner cell masses of mouse blastocysts between days 3.5 and 5.0 post coitum, or from the inner cell masses of delayed blastocysts. However, it has been suggested that some aspects of their biology may be more similar to embryonic ectoderm from early post implantation embryos, than to pre-implantation blastocysts. It has also been suggested that human embryonic stem cells are more similar to mouse stem cells derived from early embryonic ectoderm, than to mouse ES cells. Until recently, however, it has been difficult to answer these questions definitively, as the quantity of tissue available from early embryos is extremely limiting. However, we have used two rounds of RNA amplification on early embryonic samples and individual colonies picked from ES cell culture to compare the transcriptomic profiles of mouse ES cells with the embryonic tissues from with these cells are commonly derived, and with slightly later embryonic stages. We have found that ES cells are notably most similar in profile to embryonic ectoderm at day 5.5 (EE5.5) . ES cells cannot be derived from this post implantation embryonic tissue, so this result is in some ways surprising. However, ES cells show key differences from EE5.5, including the expression of markers of pluripotency such as Oct4 and Nanog (see gene lists acompanying this record), which explain how their ability to contribute to all tissues of the embryo proper is retained. Keywords: cell type comparison

Project description:An important facet of the oocyte to embryo transition in mammals is the rapid elimination of a subset of the maternal products that got accumulated during oogenesis. RNA studies support a view that the transition occurs quite rapidly. Whether this applies also for proteins remains to be determined beyond the very few cases known to date. We report that COPS3, the 3rd subunit of the COP9 signalosome complex, forms a large protein deposit in mouse oocytes (94th percentile of the riBAQ distribution). The one and only knock-out study had previously shown that Cops3 null (-/-) mouse embryos obtained from heterozygous intercrosses arrested after 5.5 dpc and were resorbed by 8.5 dpc mainly due to increased cell death in the epiblast (PMID: 12972600). However, more recent studies from our group ascribed Cops3 gene with a developmental role already at the 2-cell stage. Specifically, the sister blastomeres differ from each other in Cops3 mRNA levels and distribution, preceding the discordance of epiblast formation in derivative twin blastocysts, and implicating Cops3 in the molecular underpinnings of totipotency. This discrepancy between original knock-out result and recent observations can be resolved if we posit that the first requirement for Cops3 gene function was bridged by maternal protein that outlived the locus excision, consistent with the observation that Cops3 -/- blastocysts had the same immunostaining intensity as the +/- or +/+ counterparts in the original knock-out study. Thus, these contradicting observations support a hypothesis that 5.5 dpc may not have been the first time when the gene function was needed in development, but the second time. To test this hypothesis, pronuclear-stage mouse oocytes were subjected to an immunological method that directly targets the protein of interest by way of proteasomal degradation of the COPS3-antibody-TRIM21 complex. Briefly, pronuclear-stage mouse oocytes were microinjected with a mixture of the COPS3-specific purified monoclonal IgG antibody, mCherry-Trim21 mRNA and Oregon Green dextran beads (inert tracer). Injected oocytes were then allowed to develop and collected for lysis 24 hours later, when they reached the 2-cell stage but were not able to progress further. In addition to the COPS3, we targeted two additional proteins, OCT4 and TEAD4. The following seven experimental groups were examined by liquid chromatography-mass spectrometry (LC-MS/MS) using the pipeline described previously by Israel et al.: 1) non-manipulated 2-cell embryos; 2) 2-cell embryos from oocytes injected with Oregon Green dextran beads; 3) 2-cell embryos from oocytes injected with mCherry-Trim21 mRNA and Oregon Green; 4) 2-cell embryos from oocytes injected with mCherry-Trim21 mRNA, Oregon Green and anti-COPS3 (Abcam 79698); 5) 2-cell embryos from oocytes injected with anti-COPS3 (Abcam 79698) and Oregon Green; 6) 2-cell embryos from oocytes injected with mCherry-Trim21 mRNA, Oregon Green and anti-TEAD4 (Abcam 58310); 7) 2-cell embryos from oocytes injected with mCherry-Trim21 mRNA, Oregon Green and anti-OCT4 (Abcam 181557).

Dataset Information

DIS3L, cytoplasmic exosome catalytic subunit, is essential for development but not cell viability in mice.

Dataset's files

Publications

DIS3L, cytoplasmic exosome catalytic subunit, is essential for development but not cell viability in mice.

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