Retrospective analysis: reproducibility of interblastomere differences of mRNA expression in 2-cell stage mouse embryos is remarkably poor due to combinatorial mechanisms of blastomere diversification

Casser, Ellen; Israel, Steffen; Schlatt, Stefan; Nordhoff, Verena; Boiani, Michele

doi:10.1093/molehr/gay021

Cited by 5 publications

(8 citation statements)

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“…, 1998; Casser et al. , 2017, 2018). The singletons had been dismissed historically as exceptions to the rule, experimental errors or idiosyncrasies.…”

Section: Introductionmentioning

confidence: 99%

Differences in blastomere totipotency in 2-cell mouse embryos are a maternal trait mediated by asymmetric mRNA distribution

Casser

Wdowik

Israel

et al. 2019

Molecular Human Reproduction

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It is widely held that the first two blastomeres of mammalian embryos are equally totipotent and that this totipotency belongs to the group of regulative properties. However, this interpretation neglects an important aspect: evidence only came from successful monozygotic twins which can speak only for those pairs of half-embryos that are able to regulate in the first place. Are the frequently occurring incomplete pairs simply an artefact, or do they represent a real difference, be it in the imperfect blastomere’s ability to regulate growth or in the distribution of any compound X that constrains regulation? Using the model system of mouse embryos bisected at the 2-cell stage after fertilization, we present evidence that the interblastomere differences evade regulation by external factors and are already latent in oocytes. Specifically, an interblastomere imbalance of epiblast production persists under the most diverse culture conditions and applies to the same extent in parthenogenetic counterparts. As a result, cases in which twin blastocysts continued to develop in only one member account for 65 and 57% of zygotic and parthenogenetic pairs, respectively. The interblastomere imbalance is related to the subcellular distribution of gene products, as documented for the epiblast-related gene Cops3, using mRNA FISH in super-resolution mode confocal microscopy. Blastomere patterns of Cops3 mRNA distribution are α-amanitin-resistant. Thus, the imbalance originates not from de novo transcription, but from influences which are effective before fertilisation. These data expose previously unrecognized limits of regulative capacities of 2-cell stage blastomeres and point to aspects of cytoplasmic organization of the mouse oocyte that segregate unequally to blastomeres during cleavage.

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“…, 1998; Casser et al. , 2017, 2018). The singletons had been dismissed historically as exceptions to the rule, experimental errors or idiosyncrasies.…”

Section: Introductionmentioning

confidence: 99%

Differences in blastomere totipotency in 2-cell mouse embryos are a maternal trait mediated by asymmetric mRNA distribution

Casser

Wdowik

Israel

et al. 2019

Molecular Human Reproduction

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“…Thus, we should not expect transcripts to embody all of the interblastomere variability. Secondly, housekeeping genes, which totipotency might also rely on, were excluded from our analysis (Casser et al 2018). Thirdly, irreproducibility might be genuine: The best case in point is the very low probability of finding an egg genetically identical to another egg, given the complexity of meiotic recombination.…”

Section: In Search Of Molecular Correlates and Origins Of Unequal Blamentioning

confidence: 99%

“…The list of genes whose transcripts were used to define the two-cell-like state of ES cells and that were considered among the candidate genes for totipotency markers in mice includes: MERVL, and Eif1a-like genes, Zscan4b-Zscan4f, Zfp352, Tdpoz1-5 (Morgani & Brickman 2014, Wu et al 2017. Those genes were not part of the common differences found between blastomere at the two-cell stage (Casser et al 2018), simply because selection criteria (two-fold difference of mRNA level in at least 50% of the pairs) were not fulfilled. Functionally, two cell-like stem cells can contribute to both embryonic and extraembryonic lineages in chimeras; therefore, they could be called 'totipotent' according to the more permissive definition of totipotency, which is expressly not taken as the default in this review.…”

Section: In Search Of Molecular Correlates and Origins Of Unequal Blamentioning

confidence: 99%

“…In fact, new functional and molecular data produced with mouse embryos (Casser et al 2017(Casser et al , 2018 point to a need to reevaluate the long-held assumption of equal totipotency in two-cell stage blastomeres. This will be the goal of the second part of this review, which will reexamine the original data on the reproductive totipotency of single blastomeres from various mammals (mouse, rat, cow, sheep, goat and monkey) and from other organisms when relevant.…”

Section: Introductionmentioning

confidence: 99%

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Totipotency continuity from zygote to early blastomeres: a model under revision

et al. 2019

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The mammalian zygote is a totipotent cell that generates all the cells of a new organism through embryonic development. However, if one asks about the totipotency of blastomeres after one or two zygotic divisions, opinions differ. As it is impossible to determine the individual developmental potency of early blastomeres in an intact embryo, experiments of blastomere isolation were conducted in various species, showing that two-cell blastomeres could give rise to a new organism when sister cells were separated. A mainstream interpretation was that each of the sister mammalian blastomeres was equally totipotent. However, reevaluation of those experiments raised some doubts about the real prevalence of cases in which this interpretation could truly be validated. We compiled experiments that tested the individual developmental potency of early mammalian blastomeres in a cell-autonomous way (i.e. excluding nuclear transfer and chimera production). We then confronted the developmental abilities with reported molecular differences between sister blastomeres. The reevaluated observations were at odds with the mainstream view: A viable two-cell embryo can already include one non-totipotent blastomere. We were, thus, led to propose a revised model for totipotency continuity based on the construction of the zygote as a mosaic, which accounts for differential inheritance of totipotency-relevant components between sister blastomeres. This takes place with no preordained mechanisms that would ensure a reproducible partition. This model, which is compatible with the body of data on regulative properties of mammalian early embryos, aims at tempering the rigid interpretation that discounted maternal constraints on totipotency.Reproduction (2019) 158 R49-R65

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“…Current knowledge points at germline stem cell survival 15,16 , meiosis 17 and epiblast survival in the postimplantation embryo 6 , while the preimplantation embryo seems unaffected. We were, therefore, surprised to find that when 2-cell mouse embryos were bisected and followed up in development, the Cops3 mRNA inter-blastomere difference correlated, three days later, with the number of epiblast cells found in monozygotic twin blastocysts derived from the sister blastomeres 5,18,19 .…”

Section: Introductionmentioning

confidence: 97%

A Critical Role for The COP9 Signalosome Subunit 3 as A Gatekeeper of Genome Integrity in 2-Cell Mouse Embryos

Israel

Drexler

Fuellen

et al. 2021

Preprint

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Investigations of genes required in early mammalian development are complicated by protein deposits of maternal products, which continue to operate after the gene locus has been disrupted. This leads to an underestimation of the number of genes known to be needed during the embryonic phase of cellular totipotency (up to the 4-cell stage). Here, we expose a critical role of the gene Cops3 by showing that it protects genome integrity during the 2-cell stage of mouse embryo development, in contrast to the previous functional assignment at postimplantation. This new role is mediated by a large, stable and hitherto overlooked deposit of maternal protein. Since protein abundance and stability defeat prospects of DNA- or RNA-based gene inactivation, we adopted a protein strategy of gene inactivation: antigen masking or TRIM21-mediated proteasomal degradation of COPS3. Both resulted in 2-cell embryo lethality, but the expected degradation remained outstanding, because the major fraction of the total COPS3 is secluded in a submembrane cortical rim that withstands extraction with detergent, thereby exposing a soluble vs. insoluble fraction of COPS3, which we ascribe with distinct functional properties. In mechanistic terms, transcriptomic and metabolic analyses reveal that soluble COPS3 is involved in several processes, which, however, converge on DNA endoreduplication and the accumulation of DNA strand breaks in the 2-cell nucleus, where the minor soluble fraction of COPS3 is placed. Thus, we have shown the critical role of maternal protein deposits in development, and we have also highlighted the distinction between the site of accumulation (cell cortex) and point of use (nucleus), which is a dimension of the protein-based strategies of gene inactivation not yet fully appreciated.

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Retrospective analysis: reproducibility of interblastomere differences of mRNA expression in 2-cell stage mouse embryos is remarkably poor due to combinatorial mechanisms of blastomere diversification

Abstract: This study was supported by the Deutsche Forschungsgemeinschaft (grant DFG BO 2540-4-3 to M.B. and grant NO 413/3-3 to V.N.). The authors declare that they have no competing financial interests.

Cited by 5 publications

References 68 publications

Differences in blastomere totipotency in 2-cell mouse embryos are a maternal trait mediated by asymmetric mRNA distribution

Differences in blastomere totipotency in 2-cell mouse embryos are a maternal trait mediated by asymmetric mRNA distribution

Totipotency continuity from zygote to early blastomeres: a model under revision

A Critical Role for The COP9 Signalosome Subunit 3 as A Gatekeeper of Genome Integrity in 2-Cell Mouse Embryos

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