Molecular insights into reprogramming-initiation events mediated by the OSKM gene regulatory network

PLoS One. 2011;6(8):e24351. doi: 10.1371/journal.pone.0024351. Epub 2011 Aug 31.

Abstract

Somatic cells can be reprogrammed to induced pluripotent stem cells by over-expression of OCT4, SOX2, KLF4 and c-MYC (OSKM). With the aim of unveiling the early mechanisms underlying the induction of pluripotency, we have analyzed transcriptional profiles at 24, 48 and 72 hours post-transduction of OSKM into human foreskin fibroblasts. Experiments confirmed that upon viral transduction, the immediate response is innate immunity, which induces free radical generation, oxidative DNA damage, p53 activation, senescence, and apoptosis, ultimately leading to a reduction in the reprogramming efficiency. Conversely, nucleofection of OSKM plasmids does not elicit the same cellular stress, suggesting viral response as an early reprogramming roadblock. Additional initiation events include the activation of surface markers associated with pluripotency and the suppression of epithelial-to-mesenchymal transition. Furthermore, reconstruction of an OSKM interaction network highlights intermediate path nodes as candidates for improvement intervention. Overall, the results suggest three strategies to improve reprogramming efficiency employing: 1) anti-inflammatory modulation of innate immune response, 2) pre-selection of cells expressing pluripotency-associated surface antigens, 3) activation of specific interaction paths that amplify the pluripotency signal.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cellular Reprogramming / genetics*
  • Cellular Senescence / genetics
  • Epithelial-Mesenchymal Transition / genetics
  • Fibroblasts / metabolism
  • Gene Expression Regulation
  • Gene Regulatory Networks / genetics*
  • Humans
  • Kruppel-Like Factor 4
  • Kruppel-Like Transcription Factors / genetics*
  • Kruppel-Like Transcription Factors / metabolism
  • Male
  • Mice
  • Models, Biological
  • Octamer Transcription Factor-3 / genetics*
  • Octamer Transcription Factor-3 / metabolism
  • Pluripotent Stem Cells / metabolism
  • Proto-Oncogene Proteins c-myc / genetics*
  • Proto-Oncogene Proteins c-myc / metabolism
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Retroviridae / genetics
  • SOXB1 Transcription Factors / genetics*
  • SOXB1 Transcription Factors / metabolism
  • Time Factors
  • Transcription, Genetic
  • Transcriptome
  • Transduction, Genetic
  • Transfection
  • Tumor Suppressor Protein p53 / metabolism

Substances

  • KLF4 protein, human
  • Klf4 protein, mouse
  • Kruppel-Like Factor 4
  • Kruppel-Like Transcription Factors
  • Octamer Transcription Factor-3
  • Proto-Oncogene Proteins c-myc
  • RNA, Messenger
  • SOXB1 Transcription Factors
  • Tumor Suppressor Protein p53