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NOTCH signaling pathway is required for bovine early embryonic development

ABSTRACT: The NOTCH signaling pathway plays an important role in regulating various biological processes, including cell proliferation, lineage specification and apoptosis. Multiple components of the NOTCH pathway have been identified in mammalian preimplantation embryos. However, the precise role of NOTCH pathway in early embryonic development is poorly understood, especially in domestic animals. Here, we show that key transcripts of the NOTCH pathway exhibit a dynamic pattern throughout early embryonic development with an abundant level before embryonic genome activation in cattle. We also confirmed the presence of active NOTCH1 and RBPJ. By using pharmacological and RNAi tools, we demonstrated that the NOTCH pathway is required for the proper development of bovine early embryos. This functional consequence could be partly attributed to the major transcriptional mediator-RBPJ, whose deficiency also compromised the embryo quality. Indeed, we observed a significant increase of histone H3 serine 10 phosphorylation (pH3S10, a mitosis marker) positive blastomeres in not only NOTCH1 depleted embryos but RBPJ depleted ones. Importantly, RNA-seq analysis revealed that either NOTCH1 or RBPJ depletion triggers a reduction in H1FOO that encodes the oocyte-specific linker histone H1 variant. Interestingly, depleting H1FOO also results in detrimental effects on the developmental competence of early embryos, similar with NOTCH1 inhibition. Overall, our results reveal a crucial role for NOTCH pathway in regulating bovine preimplantation development, likely by controlling cell proliferation and maintaining H1FOO expression.

ORGANISM(S): Bos taurus

PROVIDER: GSE158679 | GEO | 2021/01/31

REPOSITORIES: GEO

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Project description:The signaling cascades that direct the morphological differentiation of the vascular system during early embryogenesis are not well defined. To further understand the role of Notch signaling during endothelial differentiation, this study uses both an in vivo gain-of-function and in vivo loss-of-function approach. At embryonic day 9.5, embryos with activated Notch1 signaling in the endothelia display a variety of growth and cardiovascular defects, and die soon after E10.5. Most notably, the extra-embryonic vasculature of the yolk sac displays remodeling differentiation defects. In the wild-type yolk sac, the primary vascular network has begun to reorganize, forming the large primary vessels and the smaller capillaries. In the activated Notch1 embryos remodeling is defective; the vasculature have an enlarged surface with decreased inter-vessel space. Embryos with ablated Notch signaling also display growth and vascular defects at E9.5 similar to the activated Notch1 embryos, however they exhibit a lack of vascular remodeling in the yolk sac, retaining the simple vascular plexus seen at E8.5. These results indicate that Notch signaling plays a critical role in the remodeling of the vasculature in the early embryo, particularly in the extra embryonic region. A conditional transgenic system was used in this study to activate Notch signaling. The ubiquitous ROSA26Notch transgene with a Neo/stop cassette flanked by loxP sites, followed by the N1-ICD cDNA, was recombined with a Tie2-CRE mouse, resulting in the removal of the STOP cassette and the subsequent activation of the Notch1-intracellular domain. This allowed for the overexpression and expansion of Notch signaling in all endothelial cells. Male Tie2-Cre mice were mated with female ROSA26Notch mice and resulting embryos were dissected at embryonic day 9.5. To ablate Notch signaling, Tie2-Cre mice were used in a two generation cross to obtain Tie2-Cre; Rbpj flox/flox embryos. These embryos lack RBPJ binding activity in the endothelia. In both instances embryos were used for genotyping and the yolk sac were separated and used to isolate total RNA with an RNeasy mini kit. The RNA was analyzed with the Mouse Genome 430A Array from Affymetrix. Samples were performed in duplicate, and RNA from wild type yolk sac tissues was compared to activated Notch and RBPJ loss-of-function yolk sac tissues.

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NOTCH signaling pathway is required for bovine early embryonic development

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