Optogenetic manipulation of cGMP in cells and animals by the tightly light-regulated guanylyl-cyclase opsin CyclOp

Nat Commun. 2015 Sep 8;6:8046. doi: 10.1038/ncomms9046.

Abstract

Cyclic GMP (cGMP) signalling regulates multiple biological functions through activation of protein kinase G and cyclic nucleotide-gated (CNG) channels. In sensory neurons, cGMP permits signal modulation, amplification and encoding, before depolarization. Here we implement a guanylyl cyclase rhodopsin from Blastocladiella emersonii as a new optogenetic tool (BeCyclOp), enabling rapid light-triggered cGMP increase in heterologous cells (Xenopus oocytes, HEK293T cells) and in Caenorhabditis elegans. Among five different fungal CyclOps, exhibiting unusual eight transmembrane topologies and cytosolic N-termini, BeCyclOp is the superior optogenetic tool (light/dark activity ratio: 5,000; no cAMP production; turnover (20 °C) ∼17 cGMP s(-1)). Via co-expressed CNG channels (OLF in oocytes, TAX-2/4 in C. elegans muscle), BeCyclOp photoactivation induces a rapid conductance increase and depolarization at very low light intensities. In O2/CO2 sensory neurons of C. elegans, BeCyclOp activation evokes behavioural responses consistent with their normal sensory function. BeCyclOp therefore enables precise and rapid optogenetic manipulation of cGMP levels in cells and animals.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Blastocladiella
  • Caenorhabditis elegans
  • Caenorhabditis elegans Proteins / metabolism
  • Carbon Dioxide
  • Chemoreceptor Cells / metabolism
  • Cyclic GMP / metabolism*
  • Cyclic Nucleotide-Gated Cation Channels / metabolism
  • Guanylate Cyclase / metabolism*
  • HEK293 Cells
  • Humans
  • Ion Channels / metabolism
  • Light*
  • Microscopy, Fluorescence
  • Oocytes / metabolism
  • Opsins / metabolism
  • Optical Imaging
  • Optogenetics / methods*
  • Oxygen
  • Patch-Clamp Techniques
  • Rhodopsin / metabolism*
  • Xenopus

Substances

  • Caenorhabditis elegans Proteins
  • Cyclic Nucleotide-Gated Cation Channels
  • Ion Channels
  • Opsins
  • tax-2 protein, C elegans
  • tax-4 protein, C elegans
  • Carbon Dioxide
  • Rhodopsin
  • Guanylate Cyclase
  • Cyclic GMP
  • Oxygen