Synthetic embryos complete gastrulation to neurulation and organogenesis.

Embryonic stem cells (ESC) can undergo many aspects of mammalian embryogenesis in vitro1-5, but their developmental potential is substantially extended by interactions with extraembryonic stem cells, including trophoblast stem cells (TSCs), extraembryonic endoderm stem cells (XEN), and inducible-XEN cells (iXEN)6-11. Here, we assembled stem-cell derived embryos in vitro from mouse ESCs, TSCs and iXEN cells and showed that they recapitulate whole natural mouse embryo development in utero to day 8.5. Our embryo model displays head-folds with defined forebrain and midbrain regions and develops a beating heart-like structure, a trunk comprising a neural tube and somites, a tail bud containing neuromesodermal progenitors, a gut tube, and primordial germ cells. This complete embryo model develops within an extra-embryonic yolk sac that initiates blood island development. Importantly, we demonstrate that the neurulating embryo model assembled from Pax6 knockout-ESCs aggregated with wild-type TSCs and iXENs recapitulates the ventral domain expansion of the neural tube that occurs in natural, ubiquitous Pax6 knockout embryos. Thus, these complete embryoids are a powerful in vitro model for dissecting the roles of diverse lineages and genes in development. Our results demonstrate the self-organization ability of embryonic and two types of extra-embryonic stem cells to reconstitute mammalian development through and beyond gastrulation to neurulation and early organogenesis.NIH Pioneer Award (DP1 HD104575-01), European Research Council (669198), the Wellcome Trust (207415/Z/17/Z), Open Philanthropy/Silicon Valley Community Foundation and Weston Havens Foundation and the Centre for Trophoblast Research. FH was supported by the ERC (69566) and the Wellcome Trust (WT108438/C/15/Z); JdJ was supported by the Biotechnology and Biological Sciences Research Council. CEH was supported by the Centre for Trophoblast Research, and the Leventis Foundation. A grant from the Paul G. Allen Frontiers Group (Allen Discovery Centre for Cell Lineage Tracing) supported MZG, ME and JS. JS is also supported by the National Human Genome Research Institute (1UM1HG011586 to J.S.; R01HG010632 to J.S.) and is an Investigator of the Howard Hughes Medical Institute. HG is supported by a Biology and Biological Engineering postdoctoral fellowship from Caltech. DY is supported by a NIH-NRSA postdoctoral fellowship (5F32HD105442

Embryo model completes gastrulation to neurulation and organogenesis.

Embryonic stem (ES) cells can undergo many aspects of mammalian embryogenesis in vitro1-5, but their developmental potential is substantially extended by interactions with extraembryonic stem cells, including trophoblast stem (TS) cells, extraembryonic endoderm stem (XEN) cells and inducible XEN (iXEN) cells6-11. Here we assembled stem cell-derived embryos in vitro from mouse ES cells, TS cells and iXEN cells and showed that they recapitulate the development of whole natural mouse embryo in utero up to day 8.5 post-fertilization. Our embryo model displays headfolds with defined forebrain and midbrain regions and develops a beating heart-like structure, a trunk comprising a neural tube and somites, a tail bud containing neuromesodermal progenitors, a gut tube, and primordial germ cells. This complete embryo model develops within an extraembryonic yolk sac that initiates blood island development. Notably, we demonstrate that the neurulating embryo model assembled from Pax6-knockout ES cells aggregated with wild-type TS cells and iXEN cells recapitulates the ventral domain expansion of the neural tube that occurs in natural, ubiquitous Pax6-knockout embryos. Thus, these complete embryoids are a powerful in vitro model for dissecting the roles of diverse cell lineages and genes in development. Our results demonstrate the self-organization ability of ES cells and two types of extraembryonic stem cells to reconstitute mammalian development through and beyond gastrulation to neurulation and early organogenesis.NIH Pioneer Award (DP1 HD104575-01), European Research Council (669198), the Wellcome Trust (207415/Z/17/Z), Open Philanthropy/Silicon Valley Community Foundation and Weston Havens Foundation and the Centre for Trophoblast Research. FH was supported by the ERC (69566) and the Wellcome Trust (WT108438/C/15/Z); JdJ was supported by the Biotechnology and Biological Sciences Research Council. CEH was supported by the Centre for Trophoblast Research, and the Leventis Foundation. A grant from the Paul G. Allen Frontiers Group (Allen Discovery Centre for Cell Lineage Tracing) supported MZG, ME and JS. JS is also supported by the National Human Genome Research Institute (1UM1HG011586 to J.S.; R01HG010632 to J.S.) and is an Investigator of the Howard Hughes Medical Institute. HG is supported by a Biology and Biological Engineering postdoctoral fellowship from Caltech. DY is supported by a NIH-NRSA postdoctoral fellowship (5F32HD105442

Context-dependent functional compensation between Ythdf m6A reader proteins

The N6-methyladenosine (m6A) modification is the most prevalent post-transcriptional mRNA modification, regulating mRNA decay and splicing. It plays a major role during normal development, differentiation, and disease progression. The modification is regulated by a set of writer, eraser, and reader proteins. The YTH domain family of proteins consists of three homologous m6A-binding proteins, Ythdf1, Ythdf2, and Ythdf3, which were suggested to have different cellular functions. However, their sequence similarity and their tendency to bind the same targets suggest that they may have overlapping roles. We systematically knocked out (KO) the Mettl3 writer, each of the Ythdf readers, and the three readers together (triple-KO). We then estimated the effect in vivo in mouse gametogenesis, postnatal viability, and in vitro in mouse embryonic stem cells (mESCs). In gametogenesis, Mettl3-KO severity is increased as the deletion occurs earlier in the process, and Ythdf2 has a dominant role that cannot be compensated by Ythdf1 or Ythdf3, due to differences in readers' expression pattern across different cell types, both in quantity and in spatial location. Knocking out the three readers together and systematically testing viable offspring genotypes revealed a redundancy in the readers' role during early development that is Ythdf1/2/3 gene dosage-dependent. Finally, in mESCs there is compensation between the three Ythdf reader proteins, since the resistance to differentiate and the significant effect on mRNA decay occur only in the triple-KO cells and not in the single KOs. Thus, we suggest a new model for the Ythdf readers function, in which there is profound dosage-dependent redundancy when all three readers are equivalently coexpressed in the same cell types

Neutralizing Gatad2a-Chd4-Mbd3/NuRD Complex Facilitates Deterministic Induction of Naive Pluripotency

Mbd3, a member of nucleosome remodeling and deacetylase (NuRD) co-repressor complex, was previously identified as an inhibitor for deterministic induced pluripotent stem cell (iPSC) reprogramming, where up to 100% of donor cells successfully complete the process. NuRD can assume multiple mutually exclusive conformations, and it remains unclear whether this deterministic phenotype can be attributed to a specific Mbd3/NuRD subcomplex. More-over, since complete ablation of Mbd3 blocks somatic cell proliferation, we aimed to explore functionally relevant alternative ways to neutralize Mbd3-dependent NuRD activity. We identify Gatad2a, a NuRD-specific subunit, whose complete deletion specifically disrupts Mbd3/NuRD repressive activity on the pluripotency circuitry during iPSC differentiation and reprogramming without ablating somatic cell proliferation. Inhibition of Gatad2a facilitates deterministic murine iPSC reprogramming within 8 days. We validate a distinct molecular axis, Gatad2a-Chd4-Mbd3, within Mbd3/NuRD as being critical for blocking reestablishment of naive pluripotency and further highlight signaling-dependent and post-translational modifications of Mbd3/NuRD that influence its interactions and assembly