Defining molecular cornerstones during fibroblast to iPS cell reprogramming in mouse.
Ectopic expression of the transcription factors Oct4, Sox2, c-Myc, and Klf4 in fibroblasts generates induced pluripotent stem (iPS) cells. Little is known about the nature and sequence of molecular events accompanying nuclear reprogramming. Using doxycycline-inducible vectors, we have shown that exogenous factors are required for about 10 days, after which cells enter a self-sustaining pluripotent state. We have identified markers that define cell populations prior to and during this transition period. While downregulation of Thy1 and subsequent upregulation of SSEA-1 occur at early time points, reactivation of endogenous Oct4, Sox2, telomerase, and the silent X chromosome mark late events in the reprogramming process. Cell sorting with these markers allows for a significant enrichment of cells with the potential to become iPS cells. Our results suggest that factor-induced reprogramming is a gradual process with defined intermediate cell populations that contain the majority of cells poised to become iPS cells.
Stadtfeld
Matthias
M
Massachusetts General Hospital Cancer Center and Center for Regenerative Medicine, 185 Cambridge Street, Boston, MA 02114, USA.
Maherali
Nimet
N
Breault
David T
DT
Hochedlinger
Konrad
K
D000818:Animals; D054337:Cell Dedifferentiation / Q000235:genetics; D002478:Cells, Cultured; D005260:Female; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism*; D005822:Genetic Vectors; D000090062:Kruppel-Like Factor 4; D016256:Lewis X Antigen; D051379:Mice; D008822:Mice, Transgenic; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D019098:Telomerase / Q000096:biosynthesis / Q000235:genetics; D018800:Thy-1 Antigens / Q000096:biosynthesis / Q000235:genetics; D013997:Time Factors; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014960:X Chromosome / Q000235:genetics / Q000378:metabolism
C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D016256:Lewis X Antigen; D018800:Thy-1 Antigens; D014157:Transcription Factors; D019098:Telomerase