SOCS3 deficiency drives the primed to naive pluripotency transition by sustaining STAT3 activation.

The transition between naive and primed pluripotency is governed by dynamic signaling networks and transcriptional circuits. While the janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) pathway is the master driver of naive pluripotency, the intrinsic negative feedback mechanisms that restrict its activation in primed epiblast stem cells (EpiSCs) remain incompletely defined. This study aimed to characterize the functional role of Suppressor of Cytokine Signaling 3 (SOCS3) in the primed-to-naive pluripotency transition. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas9 (CRISPR-associated protein 9) -mediated Socs3 knockout (KO) was generated in mouse EpiSCs, followed by primed-to-naive reprogramming induction in 2i/LIF [LIF (leukemia inhibitory factor), PD0325901 and CHIR99021) culture system. Molecular and phenotypic changes were evaluated via quantative real time PCR (qRT-PCR), Western blot, flow cytometry and immunofluorescence. Multilineage differentiation as-says were performed to verify pluripotency, and the STAT3-specific inhibitor Stattic was used to confirm the pathway dependence of the reprogramming phenotype. Socs3 was highly expressed in naive embryonic stem cells (ESCs) but minimally detected in EpiSCs. Socs3 deletion uncoupled the JAK/STAT3 negative feedback loop, causing sustained STAT3 Tyr705 phosphorylation that drove rapid and successful primed-to-naive conversion. The resulting reprogrammed naive ESCs (rnESCs) reactivated the core naive transcriptional network and acquired multilineage differentiation potential. Socs3 deficiency also delayed exit from naive pluripotency, and Stattic treatment completely abrogated Socs3 KO-mediated reprogramming. SOCS3 acts as a pivotal inducible barrier to the primed-to-naive pluripotency transition. Eliminating SOCS3-mediated negative regulation to sustain STAT3 activation is an effective strategy to overcome stem cell reprogramming barriers, providing a key target for the precise manipulation of pluripotent stem cell (PSC) fate.