Reprogramming of murine and human somatic cells using a single polycistronic vector

Directed reprogramming of somatic cells by defined factors provides a novel method for the generation of patient-specific stem cells with the potential to bypass both the practical and ethical concerns associated with somatic cell nuclear transfer (SCNT) and human embryonic stem (hES) cells. Althoug...

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Veröffentlicht in:Proceedings of the National Academy of Sciences - PNAS Jg. 106; H. 1; S. 157
Hauptverfasser: Carey, Bryce W, Markoulaki, Styliani, Hanna, Jacob, Saha, Kris, Gao, Qing, Mitalipova, Maisam, Jaenisch, Rudolf
Format: Journal Article
Sprache:Englisch
Veröffentlicht: United States 06.01.2009
Schlagworte:
Animals
Cells
Cells, Cultured
Cellular Reprogramming - genetics
Fibroblasts - cytology
Genetic Vectors - genetics
Keratinocytes - cytology
Kruppel-Like Transcription Factors - genetics
Mice
Octamer Transcription Factor-3 - genetics
Pluripotent Stem Cells - cytology
Proto-Oncogene Proteins c-myc - genetics
SOXB1 Transcription Factors - genetics
Transfection - methods
Transgenes - genetics
ISSN:1091-6490, 1091-6490
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Zusammenfassung:Directed reprogramming of somatic cells by defined factors provides a novel method for the generation of patient-specific stem cells with the potential to bypass both the practical and ethical concerns associated with somatic cell nuclear transfer (SCNT) and human embryonic stem (hES) cells. Although the generation of induced pluripotent stem (iPS) cells has proven a robust technology in mouse and human, a major impediment to the use of iPS cells for therapeutic purposes has been the viral-based delivery of the reprogramming factors because multiple proviral integrations pose the danger of insertional mutagenesis. Here we report a novel approach to reduce the number of viruses necessary to reprogram somatic cells by delivering reprogramming factors in a single virus using 2A "self-cleaving" peptides, which support efficient polycistronic expression from a single promoter. We find that up to four reprogramming factors (Oct4, Sox2, Klf4, and c-Myc) can be expressed from a single virus to generate iPS cells in both embryonic and adult somatic mouse cells and we show that a single proviral copy is sufficient to generate iPS cells from mouse embryonic fibroblasts. In addition we have generated human induced pluripotent stem (hiPS) cell lines from human keratinocytes, demonstrating that a single polycistronic virus can reprogram human somatic cells.
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ISSN:1091-6490
1091-6490
DOI:10.1073/pnas.0811426106