ORIGINAL RESEARCH article
Front. Immunol.
Sec. NK and Innate Lymphoid Cell Biology
Volume 16 - 2025 | doi: 10.3389/fimmu.2025.1607664
RAG recombinase expression discriminates the development of natural killer cells
Provisionally accepted
Jasmin Sprissler1,2
Ulrich Pannicke3
Eva-Maria Rump3
Hubert Schrezenmeier3,4,5
Nicolas Casadei6,7
Michaela Pogoda6,7Laurence Kuhlburger8Morgana Barroso Oquendo8Stefan Czemmel8
Klaus-Michael Debatin2,5Miriam Erlacher2Klaus Schwarz3,4,5
Kerstin Felgentreff2,5*- 1International Graduate School in Molecular Medicine Ulm, Ulm University, Ulm, Germany
- 2Department of Pediatrics and Adolescent Medicine, University Ulm Medical Center, Ulm, Germany
- 3Institute for Transfusion Medicine, Ulm University, Ulm, Germany
- 4Institute for Clinical Transfusion Medicine and Immunogenetics Ulm, German Red Cross Blood Transfusion Service, Ulm, Germany
- 5German Center for Child and Adolescent Health (DZKJ), partner site Ulm, Ulm, Germany
- 6Institute of Medical Genetics and Applied Genomics, University Hospital and Faculty of Medicine, University of Tübingen, Tuebingen, Germany
- 7NGS Competence Center, Tübingen University Hospital, Tübingen, Baden-Württemberg, Germany
- 8Quantitative Biology Center (QBiC), University of Tübingen, Tübingen, Germany
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V(D)J recombination initiated by recombination-activating gene (RAG) endonucleases is a crucial process for the generation of diversified antigen receptors of T and B lymphocytes but regarded dispensable for innate natural killer (NK) lymphocytes lacking clonotypic receptors.To explore the impact of potential rearrangements on NK cell maturation, RAG-fate mapping reporter human induced pluripotent stem cell (iPSC) lines were generated by introduction of RSS-invEGFP constructs into the AAVS1 locus using CRISPR/Cas9 and differentiated into NK cells in vitro.GFP expression was observed in up to 14% of mature NK cells characterized by a CD45 dim CD56 dim CD57 + NKG2C +/-KIR +/-phenotype and unproductive genetic rearrangements in the IGH locus. Advanced maturation was further revealed by transcriptomic studies using RNA sequencing. Despite their strong effector function, DNA damage response and survival to ionizing radiation were compromised.These findings suggest a role of RAG expression in NK-cell ontogeny supporting the development of a terminally differentiated effector population.
Keywords: RAG-fate reporter, Induced Pluripotent Stem Cells, NK-cell differentiation, V(D)J Recombination, DNA damage response, Lymphoid Progenitor Cells, RNA sequencing
Received: 07 Apr 2025; Accepted: 20 Jun 2025.
Copyright: © 2025 Sprissler, Pannicke, Rump, Schrezenmeier, Casadei, Pogoda, Kuhlburger, Oquendo, Czemmel, Debatin, Erlacher, Schwarz and Felgentreff. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) or licensor are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
* Correspondence: Kerstin Felgentreff, Department of Pediatrics and Adolescent Medicine, University Ulm Medical Center, Ulm, Germany
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