# YTHDF2 suppresses the 2C-like state in mouse embryonic stem cells via the DUX-ZSCAN4 molecular circuit

## TL;DR

This study shows how an RNA modification reader, YTHDF2, suppresses a 2-cell-like state in mouse embryonic stem cells through interactions with a key molecular circuit.

## Contribution

The study identifies YTHDF2 as a novel regulator of the 2C-like state via m6A modification and the DUX-ZSCAN4 circuit.

## Key findings

- YTHDF2 depletion increases 2C-signature gene expression and promotes transition to 2CLCs.
- YTHDF2 binds and degrades m6A-modified 2C transcripts.
- YTHDF2 interacts with CNOT1 to suppress the 2C-like program.

## Abstract

Mouse embryonic stem cells (ESCs) consist of a rare population of heterogeneous 2-cell-like cells (2CLCs). These cells transiently recapitulate the transcriptional and epigenetic features of the 2-cell embryos, serving as a unique model for studying totipotency acquisition and embryonic development. Accumulating evidence has demonstrated that transcription factors and epigenetic modifications exert crucial functions in the transition of ESCs to 2CLCs. However, the roles of RNA modification in the regulation of the 2C-like state remain elusive. Using a DUX-induced 2CLCs system, we examine N6-methyladenosine (m6A) modification landscape transcriptome-wide and observe dynamic regulation of m6A during DUX-driven 2C-like reprogramming. Notably, many core 2C transcripts like Dux and Zscan4 are highly methylated. We identify the m6A reader protein YTHDF2 as a critical regulator of 2C-like state. Depletion of YTHDF2 facilitates robust expression of 2C-signature genes and ESCs-to-2CLCs transition. Intriguingly, YTHDF2 binds to a subset of m6A-modified 2C transcripts and promotes their decay. We further demonstrate that YTHDF2 suppresses the 2C-like program in a manner that is dependent on both m6A and the DUX-ZSCAN4 molecular circuit. Mechanistically, YTHDF2 interacts with CNOT1, a key component of the RNA deadenylase complex. Consistently, silencing of CNOT1 upregulates the 2C program and promotes ESCs-to-2CLCs transition. Collectively, our findings reveal novel insights into the epitranscriptomic regulation of the 2C-like state in mouse ESCs.