Regulation of endogenous gene expression with a small-molecule dimerizer

Nat Biotechnol. 2002 Jul;20(7):729-33. doi: 10.1038/nbt0702-729.

Abstract

Artificial transcription factors containing designer zinc-finger DNA-binding domains (DBDs) have been used to activate or repress expression of a growing number of endogenous genes. We have combined targeted zinc-finger DBD technology with a dimerizer-regulated gene expression system to permit the small-molecule control of endogenous gene transcription. We constructed a dimerizer-responsive transcription factor that incorporates an artificial zinc-finger DBD targeted to the promoter of the human VEGF gene. Introduction of this activator into human cells allowed expression of the chromosomal VEGF gene to be induced by a small-molecule dimerizer compound consisting of a nonimmunosuppressive rapamycin analog. We found that by directly regulating zinc-finger protein (ZFP) activity, we could circumvent difficulties encountered in the generation of cell lines stably expressing conventional unregulated activators. Dimerizer-dependent VEGF induction was rapid, tight, and dose dependent, and resulted in VEGF protein expression levels several-fold greater than those produced by the natural hypoxic response.

MeSH terms

  • Cell Line
  • Dimerization
  • Dose-Response Relationship, Drug
  • Endothelial Growth Factors / biosynthesis
  • Endothelial Growth Factors / genetics*
  • Gene Expression Regulation / drug effects*
  • Humans
  • Intercellular Signaling Peptides and Proteins / biosynthesis
  • Intercellular Signaling Peptides and Proteins / genetics*
  • Kidney / embryology
  • Lymphokines / biosynthesis
  • Lymphokines / genetics*
  • Models, Genetic
  • Plasmids
  • Sirolimus / administration & dosage*
  • Transcription Factors / genetics*
  • Transcription, Genetic*
  • Vascular Endothelial Growth Factor A
  • Vascular Endothelial Growth Factors
  • Zinc Fingers / genetics*

Substances

  • Endothelial Growth Factors
  • Intercellular Signaling Peptides and Proteins
  • Lymphokines
  • Transcription Factors
  • Vascular Endothelial Growth Factor A
  • Vascular Endothelial Growth Factors
  • Sirolimus