Retinoic acid signaling is critical during the totipotency window in early mammalian development

Ane Iturbide,Kamyar Hadian,Antonio Scialdone,Elias R Ruiz-Morales,Ina Rothenaigner,Gabriele Lubatti,Maria-Elena Torres-Padilla,Kenji Schorpp,Mayra L Ruiz Tejada Segura,Ahmed Agami,Camille Noll

doi:10.1038/s41594-021-00590-w

Abstract

Totipotent cells hold enormous potential for regenerative medicine. Thus, the development of cellular models recapitulating totipotent-like features is of paramount importance. Cells resembling the totipotent cells of early embryos arise spontaneously in mouse embryonic stem (ES) cell cultures. Such ‘2-cell-like-cells’ (2CLCs) recapitulate 2-cell-stage features and display expanded cell potential. Here, we used 2CLCs to perform a small-molecule screen to identify new pathways regulating the 2-cell-stage program. We identified retinoids as robust inducers of 2CLCs and the retinoic acid (RA)-signaling pathway as a key component of the regulatory circuitry of totipotent cells in embryos. Using single-cell RNA-seq, we reveal the transcriptional dynamics of 2CLC reprogramming and show that ES cells undergo distinct cellular trajectories in response to RA. Importantly, endogenous RA activity in early embryos is essential for zygotic genome activation and developmental progression. Overall, our data shed light on the gene regulatory networks controlling cellular plasticity and the totipotency program.

Highlights

Totipotency is the ability of a cell to give rise to a full organism[1,2] and encompasses the broadest cellular plasticity in the mammalian body
Cells resembling the blastomeres of 2-cell stage embryos, referred to as ‘2-cell-like-cells’ (2CLCs), arise spontaneously, constituting less than 1% of the cells8. 2CLCs share several features with 2-cell stage embryos, including a ‘2 C’ transcriptional program, characterized by genes expressed upon zygotic genome activation (ZGA), which occurs in late 2-cell embryos[8,9,10]
We addressed if inhibiting RARγ affects ZGA by analyzing MERVL expression— a key ZGA marker—in embryos treated with LY2955303. qPCR revealed a striking reduction in MERVL transcripts in 2-cell embryos upon RARγ inhibition (Fig. 6f)

Summary

Introduction

Totipotency is the ability of a cell to give rise to a full organism[1,2] and encompasses the broadest cellular plasticity in the mammalian body. This includes the transcription factor ZSCAN411 and retrotransposons from the MERVL family[12]. Much effort has been made towards understanding the mechanisms regulating the transcriptional program in 2CLCs and in 2-cell stage embryos[8,9,10,16,17,18,19,20,21] It is still unclear how 2CLCs arise, and the factors that activate the 2-cell program and regulate ZGA in vivo remain elusive. Identifying conditions that can robustly induce and stably maintain 2CLCs in culture can shed light into their regulatory networks and potentially uncover key factors activating the earliest developmental program in mammals

Methods

Results

Conclusion