Stoichiometric and temporal requirements of Oct4, Sox2, Klf4, and c-Myc expression for efficient human iPSC induction and differentiation Academic Article uri icon

Overview

MeSH Major

  • Genes, myc
  • Kruppel-Like Transcription Factors
  • Octamer Transcription Factor-3
  • Pluripotent Stem Cells
  • SOXB1 Transcription Factors

abstract

  • Human-induced pluripotent stem cells (hiPSCs) are generated from somatic cells by ectopic expression of the 4 reprogramming factors (RFs) Oct-4, Sox2, Klf4, and c-Myc. To better define the stoichiometric requirements and dynamic expression patterns required for successful hiPSC induction, we generated 4 bicistronic lentiviral vectors encoding the 4 RFs co-expressed with discernable fluorescent proteins. Using this system, we define the optimal stoichiometry of RF expression to be highly sensitive to Oct4 dosage, and we demonstrate the impact that variations in the relative ratios of RF expression exert on the efficiency of hiPSC induction. Monitoring of expression of each individual RF in single cells during the course of reprogramming revealed that vector silencing follows acquisition of pluripotent cell markers. Pronounced lentiviral vector silencing was a characteristic of successfully reprogrammed hiPSC clones, but lack of complete silencing did not hinder hiPSC induction, maintenance, or directed differentiation. The vector system described here presents a powerful tool for mechanistic studies of reprogramming and the optimization of hiPSC generation.

publication date

  • August 4, 2009

Research

keywords

  • Academic Article

Identity

Language

  • eng

PubMed Central ID

  • PMC2722286

Digital Object Identifier (DOI)

  • 10.1073/pnas.0904825106

PubMed ID

  • 19549847

Additional Document Info

start page

  • 12759

end page

  • 64

volume

  • 106

number

  • 31