CNOT3-Dependent mRNA Deadenylation Safeguards the Pluripotent State

Poly(A) tail length and mRNA deadenylation play important roles in gene regulation. However, how they regulate embryonic development and pluripotent cell fate is not fully understood. Here we present evidence that CNOT3-dependent mRNA deadenylation governs the pluripotent state. We show that CNOT3, a component of the Ccr4-Not deadenylase complex, is required for mouse epiblast maintenance. It is highly expressed in blastocysts and its deletion leads to peri-implantation lethality. The epiblast cells in Cnot3 deletion embryos are quickly lost during diapause and fail to outgrow in culture. Mechanistically, CNOT3 C terminus is required for its interaction with the complex and its function in embryonic stem cells (ESCs). Furthermore, Cnot3 deletion results in increases in the poly(A) tail lengths, half-lives, and steady-state levels of differentiation gene mRNAs. The half-lives of CNOT3 target mRNAs are shorter in ESCs and become longer during normal differentiation. Together, we propose that CNOT3 maintains the pluripotent state by promoting differentiation gene mRNA deadenylation and degradation, and we identify poly(A) tail-length regulation as a post-transcriptional mechanism that controls pluripotency.