Abstract
Unrepaired DNA damage during embryonic development can be potentially inherited by a large population of cells. However, the quality control mechanisms that minimize the contribution of damaged cells to developing embryos remain poorly understood. Here, we uncovered an ATR- and CHK1-mediated transcriptional response to replication stress (RS) in mouse embryonic stem cells (ESCs) that induces genes expressed in totipotent two-cell (2C) stage embryos and 2C-like cells. This response is mediated by Dux, a multicopy retrogene defining the cleavage-specific transcriptional program in placental mammals. In response to RS, DUX triggers the transcription of 2C-like markers such as murine endogenous retrovirus-like elements (MERVL) and Zscan4. This response can also be elicited by ETAA1-mediated ATR activation in the absence of RS. ATR-mediated activation of DUX requires GRSF1-dependent post-transcriptional regulation of Dux mRNA. Strikingly, activation of ATR expands ESCs fate potential by extending their contribution to both embryonic and extra-embryonic tissues. These findings define a novel ATR dependent pathway involved in maintaining genome stability in developing embryos by controlling ESCs fate in response to RS.
Highlights
embryonic stem cells (ESCs) are characterized by self-renewal and the ability to propagate for several cycles in vitro and in vivo (Giachino et al, 2013)
To understand whether the increase in the number of 2C-like cells was a response to replication stress (RS) or it was limited to APH treatment, we exposed pZscan4-Emerald ESCs generated by the stable introduction of the EmeraldGFP reporter under the Zscan4c promoter (Zalzman et al, 2010) to a range of RS-inducing agents, including APH, hydroxyurea (HU) and ultraviolet light (UV) (Cimprich and Cortez, 2008)
The increase in the number of Em+ ESCs upon short exposure to UV revealed that the continuous presence of the RS-inducing agent was not necessary for the activation of 2C-like cells (Figure 1e; Figure 1—source data 1), suggesting that replication fork stalling induced by UV-mediated DNA lesions was sufficient to trigger this pathway
Summary
ESCs are characterized by self-renewal and the ability to propagate for several cycles in vitro and in vivo (Giachino et al, 2013). Even if ESCs exhibit several markers of RS (Ahuja et al, 2016), they are able to maintain genome integrity more efficiently than differentiated cells (Giachino et al, 2013). The mechanisms underlying such distinctive feature are largely unknown. ESC colonies harbor a small transient subpopulation of cells (2C-like cells) with functional and transcriptional features of totipotent 2C-stage embryos (Choi et al, 2017; Ishiuchi et al, 2015; Macfarlan et al, 2012).
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