Oct4 redox sensitivity potentiates reprogramming and differentiation

Abstract

SUMMARYThe transcription factor Oct4/Pou5f1 is a component of the regulatory circuitry governing pluripotency and is widely used to generate induced pluripotent stem cells (iPSCs) from somatic cells. Here we use domain swapping and mutagenesis to compare Oct4’s reprogramming activity with the reprogramming-incompetent paralog Oct1/Pou2f1, identifying a redox-sensitive DNA binding domain cysteine residue (Cys48) as a key determinant of reprogramming and differentiation. In combination with the Oct4 N-terminus, mutating Oct1’s serine at this position to cysteine is sufficient to confer reprogramming activity. Conversely, Oct4C48Sreduces reprogramming potential. Oct4C48Sdesensitizes the protein to oxidative inhibition and prevents oxidation-mediated protein ubiquitylation.Pou5f1C48Spoint mutation has little effect on undifferentiated embryonic stem cells (ESCs), but upon retinoic acid (RA) treatment causes retention of Oct4 expression and deregulated differentiation.Pou5f1C48SESCs also contribute poorly to adult somatic tissues and form less differentiated teratomas. Collectively, these findings uncover a new Oct4 redox mechanism involved in both entry into and exit from pluripotency.