Rescue of DNA-PK Signaling and T-Cell Differentiation by Targeted Genome Editing in a prkdc Deficient iPSC Disease Model

Rahman, Shamim H.Kuehle, JohannesReimann, ChristianMlambo, TafadzwaAlzubi, JamalMaeder, Morgan L.Riedel, HeimoFisch, PaulCantz, TobiasRudolph, CorneliaMussolino, ClaudioJoung, J. KeithSchambach, AxelCathomen, Toni2015-06-022015Rahman, S. H., J. Kuehle, C. Reimann, T. Mlambo, J. Alzubi, M. L. Maeder, H. Riedel, et al. 2015. “Rescue of DNA-PK Signaling and T-Cell Differentiation by Targeted Genome Editing in a prkdc Deficient iPSC Disease Model.” PLoS Genetics 11 (5): e1005239. doi:10.1371/journal.pgen.1005239. http://dx.doi.org/10.1371/journal.pgen.1005239.1553-7390http://nrs.harvard.edu/urn-3:HUL.InstRepos:16121036In vitro disease modeling based on induced pluripotent stem cells (iPSCs) provides a powerful system to study cellular pathophysiology, especially in combination with targeted genome editing and protocols to differentiate iPSCs into affected cell types. In this study, we established zinc-finger nuclease-mediated genome editing in primary fibroblasts and iPSCs generated from a mouse model for radiosensitive severe combined immunodeficiency (RS-SCID), a rare disorder characterized by cellular sensitivity to radiation and the absence of lymphocytes due to impaired DNA-dependent protein kinase (DNA-PK) activity. Our results demonstrate that gene editing in RS-SCID fibroblasts rescued DNA-PK dependent signaling to overcome radiosensitivity. Furthermore, in vitro T-cell differentiation from iPSCs was employed to model the stage-specific T-cell maturation block induced by the disease causing mutation. Genetic correction of the RS-SCID iPSCs restored T-lymphocyte maturation, polyclonal V(D)J recombination of the T-cell receptor followed by successful beta-selection. In conclusion, we provide proof that iPSC-based in vitro T-cell differentiation is a valuable paradigm for SCID disease modeling, which can be utilized to investigate disorders of T-cell development and to validate gene therapy strategies for T-cell deficiencies. Moreover, this study emphasizes the significance of designer nucleases as a tool for generating isogenic disease models and their future role in producing autologous, genetically corrected transplants for various clinical applications.en-USRescue of DNA-PK Signaling and T-Cell Differentiation by Targeted Genome Editing in a prkdc Deficient iPSC Disease ModelJournal Article2015-06-0210.1371/journal.pgen.1005239