The artificial 4-zinc-finger protein Bagly binds human utrophin promoter A at the endogenous chromosomal site and activates transcription

Biochem Cell Biol. 2007 Jun;85(3):358-65. doi: 10.1139/o07-015.

Abstract

Our aim is to upregulate the expression of the dystrophin-related gene utrophin in Duchenne muscular dystrophy, in this way complementing the lack of dystrophin function. To achieve utrophin upregulation, we designed and engineered synthetic zinc-finger based transcription factors. We have previously shown that the artificial 3-zinc-finger protein Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from utrophin promoter A. Here we report a novel artificial 4-zinc-finger protein, Bagly, which binds with optimized affinity-specificity to a 12 bp DNA target sequence that is internal to human utrophin promoter A. Bagly was generated adding to Jazz protein an extra-fourth zinc finger, derived from transcription factor YY1. Importantly, the Bagly DNA target sequence is statistically present in the human genome only 210 times, about 60 fewer times than the 9 bp Jazz DNA target sequence. Thanks to its additional zinc-finger domain, Bagly protein shows enhanced transcriptional activity. Moreover, we demonstrated Bagly's effective access and binding to active chromatin in the chromosomal context and its ability to upregulate endogenous utrophin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Binding Sites / genetics
  • DNA Probes / genetics
  • HeLa Cells
  • Humans
  • Molecular Sequence Data
  • Promoter Regions, Genetic*
  • Protein Binding
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Transcriptional Activation
  • Transfection
  • Utrophin / genetics*
  • Zinc Fingers

Substances

  • DNA Probes
  • Recombinant Proteins
  • UTRN protein, human
  • Utrophin