Identification of a cell-penetrating peptide applicable to a protein-based transcription activator-like effector expression system for cell engineering

Biomaterials. 2018 Aug;173:11-21. doi: 10.1016/j.biomaterials.2018.04.040. Epub 2018 Apr 25.

Abstract

Cellular reprogramming is a promising technology in regenerative medicine, but most studies have been performed by using expression vectors. For future clinical applications, it is necessary to establish a system in which cell engineering can be manipulated without any risk of damaging the genome. Here, we identified a cell-penetrating peptide composed of 10 amino acids (RIFIHFRIGC) with nuclear trafficking activity and found that it was significantly more potent than a Tat-derived peptide or polyarginine peptide (R11). We named the peptide "nuclear trafficking peptide" (NTP) and applied it to a protein-based artificial transcription factor (NTP-ATF), which was composed of a transcription activator-like effector and transcription domain (VP64). An NTP-ATF designed to the proximal promoter region of the microRNA-302/367 cluster efficiently induced endogenous RNA expression at an extremely low concentration (0.25 nM), and repetitive treatment of mouse embryonic fibroblasts with NTP-ATF generated induced pluripotent stem-like cells, which gave chimeric mice. Together with the observation that recombinant NTP-ATF protein did not induce any apparent cytotoxicity, we propose that NTP-ATF is a promising system for cellular reprogramming applicable to regenerative medicine.

Keywords: Artificial transcription factor; Cell-penetrating peptide (CPP); Induced pluripotent stem cell (iPSC); Mouse embryonic fibroblast (MEF); Transcription activator-like effector (TALE).

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Active Transport, Cell Nucleus
  • Animals
  • Cell Engineering / methods*
  • Cell Line, Tumor
  • Cell Nucleus / metabolism
  • Cell Survival
  • Cell-Penetrating Peptides / genetics
  • Cell-Penetrating Peptides / metabolism*
  • Cell-Penetrating Peptides / pharmacology
  • Cellular Reprogramming
  • Chimera
  • Fibroblasts / cytology
  • Fibroblasts / drug effects
  • Fibroblasts / metabolism
  • Humans
  • Induced Pluripotent Stem Cells / cytology
  • Induced Pluripotent Stem Cells / drug effects
  • Induced Pluripotent Stem Cells / metabolism
  • Mice
  • MicroRNAs / genetics
  • MicroRNAs / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • Recombinant Proteins / pharmacology
  • Transcription Activator-Like Effectors / genetics
  • Transcription Activator-Like Effectors / metabolism*
  • Transcription Factors / genetics
  • Transcription Factors / metabolism

Substances

  • Cell-Penetrating Peptides
  • MicroRNAs
  • Recombinant Proteins
  • Transcription Activator-Like Effectors
  • Transcription Factors