title: Fine-tuning of iPSC derivation by an inducible reprogramming system at the protein level.
abstract: Induced pluripotent stem cells (iPSCs) generated from somatic cells by ectopic expression of reprogramming factors, e.g., POU5F1 (OCT4), KLF4, and SOX2, have great potential for regenerative medicine. However, before they can be used in a clinical setting, the mechanism of reprogramming needs to be better understood. Here, by engineering reprogramming factors to a destabilizing protein domain, we achieved inducible generation of mouse and pig iPSCs. Stability of the fusion protein was precisely regulated by the addition of the cell-permeable small molecule trimethoprim (TMP) in a dose-dependent manner. With these tools, we found that during the early and middle stages of reprogramming, exogenous OCT4 or KLF4 could be omitted, whereas exogenous SOX2 expression at early and middle stages was required for successful reprogramming. Our TMP reprogramming system is useful for defining the stoichiometry and temporal requirements of transcription factors for reprogramming.
authors.affiliation: State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Dandan
authors.initials: D
authors.lastname: Sui
authors.affiliation: State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Zhaolin
authors.initials: Z
authors.lastname: Sun
authors.affiliation: State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Chunlong
authors.initials: C
authors.lastname: Xu
authors.affiliation: Department of Human Genetics, University of Utah, Salt Lake City, UT 84112, USA.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Wu
authors.affiliation: Howard Hughes Medical Institute, University of Utah, Salt Lake City, UT 84112, USA ; Department of Human Genetics, University of Utah, Salt Lake City, UT 84112, USA.
authors.forename: Mario R
authors.initials: MR
authors.lastname: Capecchi
authors.affiliation: State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Sen
authors.initials: S
authors.lastname: Wu
authors.affiliation: State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, PR China.
authors.forename: Ning
authors.initials: N
authors.lastname: Li
mesh_terms: D000818:Animals; D002460:Cell Line; D065150:Cellular Reprogramming / Q000187:drug effects; D058750:Epithelial-Mesenchymal Transition; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism*; D013552:Swine; D014295:Trimethoprim / Q000494:pharmacology
chemical_list: C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D050814:Octamer Transcription Factor-3; D012333:RNA, Messenger; D055748:SOXB1 Transcription Factors; D014295:Trimethoprim