A gene-fusion strategy for stoichiometric and co-localized expression of light-gated membrane proteins

Nat Methods. 2011 Nov 6;8(12):1083-8. doi: 10.1038/nmeth.1766.


The precise co-localization and stoichiometric expression of two different light-gated membrane proteins can vastly improve the physiological usefulness of optogenetics for the modulation of cell excitability with light. Here we present a gene-fusion strategy for the stable 1:1 expression of any two microbial rhodopsins in a single polypeptide chain. By joining the excitatory channelrhodopsin-2 with the inhibitory ion pumps halorhodopsin or bacteriorhodopsin, we demonstrate light-regulated quantitative bi-directional control of the membrane potential in HEK293 cells and neurons in vitro. We also present synergistic rhodopsin combinations of channelrhodopsin-2 with Volvox carteri channelrhodopsin-1 or slow channelrhodopsin-2 mutants, to achieve enhanced spectral or kinetic properties, respectively. Finally, we demonstrate the utility of our fusion strategy to determine ion-turnovers of as yet uncharacterized rhodopsins, exemplified for archaerhodopsin and CatCh, or to correct pump cycles, exemplified for halorhodopsin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Artificial Gene Fusion*
  • Bacteriorhodopsins / analysis
  • Bacteriorhodopsins / biosynthesis
  • Bacteriorhodopsins / genetics
  • HEK293 Cells
  • Hippocampus / cytology
  • Humans
  • Light*
  • Molecular Sequence Data
  • Neurons / cytology
  • Neurons / metabolism
  • Rhodopsin / analysis
  • Rhodopsin / biosynthesis
  • Rhodopsin / genetics*


  • Bacteriorhodopsins
  • Rhodopsin

Associated data

  • GENBANK/JN836740
  • GENBANK/JN836741
  • GENBANK/JN836742
  • GENBANK/JN836743
  • GENBANK/JN836744
  • GENBANK/JN836745
  • GENBANK/JN836746
  • GENBANK/JN836747