METTL3 inhibits primed-to-naïve transition of pluripotent stem cells through m(6)A-YTHDF2-pluripotency/Gstp1 mRNA degradation axis.

N6-methyladenosine (m(6)A) plays crucial roles in development and cellular reprogramming. During embryonic development, pluripotency transitions from a naïve to a primed state, and modeling the reverse primed-to-naïve transition (PNT) provides a valuable framework for investigating pluripotency regulation. Here, we show that inhibiting METTL3 significantly promotes PNT in an m(6)A-dependent manner. Mechanistically, we found that suppressing METTL3 and YTHDF2 prolongs the lifetimes of pluripotency-associated mRNAs, such as Nanog and Sox2, during PNT. In addition, Gstp1 was identified as a downstream target of METTL3 inhibition and YTHDF2 knockout. Gstp1 overexpression enhances PNT, whereas its inhibition impedes the transition. Overall, our findings suggest that YTHDF2 facilitates the removal of pluripotency gene transcripts and Gstp1, thereby promoting PNT reprogramming through m(6)A-mediated posttranscriptional control.