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University in St. Louishttps://www.ebi.ac.uk/ena/browser/view/PRJNA1117755Homo sapiensxref:PubMed:39952244Naïve human pluripotent stem cells (hPSCs) model the pre-implantation epiblast. However, parent-specific epigenetic marks (imprints) are eroded in naïve hPSCs, which represents an important deviation from the epiblast in vivo. To track the dynamics of imprint erasure during naïve resetting in real time, we established a dual-colored fluorescent reporter at both alleles of the imprinted SNRPN locus. During primed-to-naïve resetting, SNRPN expression becomes biallelic in most naïve cells and biallelic SNRPN expression is irreversible upon re-priming. We utilized this live-cell reporter to evaluate chemical and genetic strategies to minimize imprint erasure. Decreasing the level of MEK/ERK inhibition or overexpressing the KRAB zinc finger protein ZFP57 protected a subset of imprints during naïve resetting. Combining these two strategies protected imprint levels to a further extent than either strategy alone. This study offers an experimental tool to track and enhance imprint stability during transitions between human pluripotent states in vitro. Overall design: Bulk RNA-seq analysis of H9 and H9-SRG primed, naïve, and alternative hESCs.ENAfalseTracking and mitigating imprint erasure during induction of naïve human pluripotency at single-cell resolution [RNA-Seq]Tracking and mitigating imprint erasure during induction of naïve human pluripotency at single-cell resolution [RNA-Seq]2025-09-242025-02-15PRJNA1117755GSE268535960639952244