Construction of Light-Activated Neurotrophin Receptors Using the Improved Light-Induced Dimerizer (iLID)

J Mol Biol. 2020 Jun 12;432(13):3739-3748. doi: 10.1016/j.jmb.2020.04.018. Epub 2020 Apr 23.

Abstract

Receptor tyrosine kinases (RTKs) play crucial roles in human health, and their misregulation is implicated in disorders ranging from neurodegenerative diseases to cancers. The highly conserved mechanism of activation of RTKs makes them especially appealing candidates for control via optogenetic dimerization methods. This work offers a strategy for using the improved light-induced dimer (iLID) system with a constructed tandem dimer of its binding partner nano (tdnano) to build light-activatable versions of RTKs. In the absence of light, the iLID-RTK is cytosolic, monomeric, and inactive. Under blue light, the iLID + tdnano system recruits two copies of iLID-RTK to tdnano, dimerizing, and activating the RTK. We demonstrate that iLID opto-iTrkA and opto-iTrkB are capable of reproducing downstream ERK and Akt signaling only in the presence of tdnano. We further show with our opto-iTrkA that the system is compatible with multi-day and population-level activation of TrkA in PC12 cells. By leveraging genetic targeting of tdnano, we achieve RTK activation at a specific subcellular location even with whole-cell illumination, allowing us to confidently probe the impact of context on signaling outcome.

Keywords: RTK; cell signaling; optogenetics; receptor tyrosine kinases.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.
  • Review

MeSH terms

  • Animals
  • Dimerization
  • Humans
  • Light
  • Nerve Growth Factors / genetics*
  • Optogenetics / trends
  • Rats
  • Receptor Protein-Tyrosine Kinases / genetics*
  • Receptors, Nerve Growth Factor / genetics*
  • Signal Transduction / genetics*
  • Signal Transduction / radiation effects

Substances

  • Nerve Growth Factors
  • Receptors, Nerve Growth Factor
  • Receptor Protein-Tyrosine Kinases