Partial promoter substitutions generating transcriptional sentinels of diverse signaling pathways in embryonic stem cells and mice

Dis Model Mech. 2012 Nov;5(6):956-66. doi: 10.1242/dmm.009696. Epub 2012 Aug 10.

Abstract

Extracellular signals in development, physiology, homeostasis and disease often act by regulating transcription. Herein we describe a general method and specific resources for determining where and when such signaling occurs in live animals and for systematically comparing the timing and extent of different signals in different cellular contexts. We used recombinase-mediated cassette exchange (RMCE) to test the effect of successively deleting conserved genomic regions of the ubiquitously active Rosa26 promoter and substituting the deleted regions for regulatory sequences that respond to diverse extracellular signals. We thereby created an allelic series of embryonic stem cells and mice, each containing a signal-responsive sentinel with different fluorescent reporters that respond with sensitivity and specificity to retinoic acids, bone morphogenic proteins, activin A, Wnts or Notch, and that can be adapted to any pathway that acts via DNA elements.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Activins / genetics
  • Activins / metabolism
  • Animals
  • Base Sequence
  • Bone Morphogenetic Proteins / genetics
  • Bone Morphogenetic Proteins / metabolism
  • Embryo, Mammalian / drug effects
  • Embryo, Mammalian / metabolism
  • Embryonic Stem Cells / drug effects
  • Embryonic Stem Cells / metabolism*
  • Genetic Engineering
  • Genetic Loci / genetics
  • Humans
  • Mice
  • Molecular Sequence Data
  • Mutation / genetics*
  • Promoter Regions, Genetic*
  • Proteins / genetics
  • RNA, Untranslated
  • Rats
  • Receptors, Notch / genetics
  • Receptors, Notch / metabolism
  • Recombination, Genetic / genetics
  • Response Elements / genetics
  • Sequence Deletion / genetics
  • Signal Transduction / drug effects
  • Signal Transduction / genetics*
  • Transcription, Genetic* / drug effects
  • Tretinoin / pharmacology
  • Wnt Signaling Pathway / drug effects
  • Wnt Signaling Pathway / genetics

Substances

  • Bone Morphogenetic Proteins
  • Gt(ROSA)26Sor non-coding RNA, mouse
  • Proteins
  • RNA, Untranslated
  • Receptors, Notch
  • activin A
  • Activins
  • Tretinoin