Reversible optogenetic control of kinase activity during differentiation and embryonic development

Vishnu V. Krishnamurthy, John S. Khamo, Wenyan Mei, Aurora J. Turgeon, Humza M. Ashraf, Payel Mondal, Dil B. Patel, Noah Risner, Ellen E. Cho, Jing Yang, Kai Zhang

Research output: Contribution to journalArticlepeer-review

Abstract

A limited number of signaling pathways are repeatedly used to regulate a wide variety of processes during development and differentiation. The lack of tools to manipulate signaling pathways dynamically in space and time has been a major technical challenge for biologists. Optogenetic techniques, which utilize light to control protein functions in a reversible fashion, hold promise for modulating intracellular signaling networks with high spatial and temporal resolution. Applications of optogenetics in multicellular organisms, however, have not been widely reported. Here, we create an optimized bicistronic optogenetic system using Arabidopsis thaliana cryptochrome 2 (CRY2) protein and the N-terminal domain of cryptochrome-interacting basic-helix-loop-helix (CIBN). In a proofof- principle study, we develop an optogenetic Raf kinase that allows reversible light-controlled activation of the Raf/MEK/ERK signaling cascade. In PC12 cells, this system significantly improves lightinduced cell differentiation compared with co-transfection. When applied to Xenopus embryos, this system enables blue lightdependent reversible Raf activation at any desired developmental stage in specific cell lineages. Our system offers a powerful optogenetic tool suitable for manipulation of signaling pathways with high spatial and temporal resolution in a wide range of experimental settings.

Original languageEnglish (US)
Pages (from-to)4085-4094
Number of pages10
JournalDevelopment (Cambridge)
Volume143
Issue number21
DOIs
StatePublished - Nov 1 2016

Keywords

  • Bicistronic gene expression
  • Cryptochrome
  • Differentiation
  • Light-controlled protein-protein interaction
  • Optogenetics
  • Xenopus

ASJC Scopus subject areas

  • Molecular Biology
  • Developmental Biology

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