Tracking and mitigating imprint erasure during induction of naive human pluripotency at single-cell resolution.

Sabine Dietmann, Monica Reyes, Thorold W Theunissen, Kyoung-Mi Park, Harald Jüppner, Ting Wang, Laura A Fischer, Brittany Meyer, Joseph E Zemke, Jessica K Harrison

Journal: Stem cell reports 2025;20(3):102419

PMID: 39952244

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Abstract

Naive human pluripotent stem cells (hPSCs) model the pre-implantation epiblast. However, parent-specific epigenetic marks (imprints) are eroded in naive hPSCs, which represents an important deviation from the epiblast in vivo. To track the dynamics of imprint erasure during naive resetting in real time, we established a dual-colored fluorescent reporter at both alleles of the imprinted SNRPN locus. During primed-to-naive resetting, SNRPN expression becomes biallelic in most naive 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 naive 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.

Address: Department of Developmental Biology and Center of Regenerative Medicine, Washington University School of Medicine, St. Louis, MO, USA.; Endocrine Unit, Department of Medicine and Pediatric Nephrology Unit, Department of Pediatrics, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.; Department of Genetics, The Edison Family Center for Genome Sciences & Systems Biology, Washington University School of Medicine, St. Louis, MO, USA.; Department of Developmental Biology and Center of Regenerative Medicine, Washington University School of Medicine, St. Louis, MO, USA; Institute for Informatics (I(2)), Washington University School of Medicine, St. Louis, MO, USA.; Department of Developmental Biology and Center of Regenerative Medicine, Washington University School of Medicine, St. Louis, MO, USA. Electronic address: [email protected].

DOI: 10.1016/j.stemcr.2025.102419

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MeSH Terms: Humans, Pluripotent Stem Cells, Single-Cell Analysis, Genomic Imprinting, Cell Differentiation, Epigenesis, Genetic, Alleles