biostudies-literatureShen ZNational Institutes of HealthNational Institute of General Medical SciencesNIGMS NIH HHS308-321https://www.ebi.ac.uk/biostudies/studies/S-EPMC11146590biostudies-literatureUnknown38(7-8)The transcription factor Oct4/Pou5f1 is a component of the regulatory circuitry governing pluripotency and is widely used to induce pluripotency from somatic cells. Here we used domain swapping and mutagenesis to study Oct4's reprogramming ability, identifying a redox-sensitive DNA binding domain, cysteine residue (Cys48), as a key determinant of reprogramming and differentiation. Oct4 Cys48 sensitizes the protein to oxidative inhibition of DNA binding activity and promotes oxidation-mediated protein ubiquitylation. <i>Pou5f1</i> ^<i>C48S</i> point mutation has little effect on undifferentiated embryonic stem cells (ESCs) but upon retinoic acid (RA) treatment causes retention of Oct4 expression, deregulated gene expression, and aberrant differentiation. <i>Pou5f1</i> <sup><i>C48SGenes & developmentOct4 redox sensitivity potentiates reprogramming and differentiation.PMC11146590R01 GM122778R01GM122778R01 GM127783Cairns BRShen ZTantin DChandrasekharan MBEvason KJManna AYi CWu YfalseOct4 redox sensitivity potentiates reprogramming and differentiation.The transcription factor Oct4/Pou5f1 is a component of the regulatory circuitry governing pluripotency and is widely used to induce pluripotency from somatic cells. Here we used domain swapping and mutagenesis to study Oct4's reprogramming ability, identifying a redox-sensitive DNA binding domain, cysteine residue (Cys48), as a key determinant of reprogramming and differentiation. Oct4 Cys48 sensitizes the protein to oxidative inhibition of DNA binding activity and promotes oxidation-mediated protein ubiquitylation. <i>Pou5f1</i> ^<i>C48S</i> point mutation has little effect on undifferentiated embryonic stem cells (ESCs) but upon retinoic acid (RA) treatment causes retention of Oct4 expression, deregulated gene expression, and aberrant differentiation. <i>Pou5f1</i> <sup><i>C48S2024-01-01T00:00:00Z2024 May2026-06-02T03:54:58.497Z2025-04-26T19:34:12.483ZS-EPMC111465903871954110.1101/gad.351411.123