Green-to-Red Photoconversion of GCaMP

PLoS One. 2015 Sep 18;10(9):e0138127. doi: 10.1371/journal.pone.0138127. eCollection 2015.

Abstract

Genetically encoded calcium indicators (GECIs) permit imaging intracellular calcium transients. Among GECIs, the GFP-based GCaMPs are the most widely used because of their high sensitivity and rapid response to changes in intracellular calcium concentrations. Here we report that the fluorescence of GCaMPs--including GCaMP3, GCaMP5 and GCaMP6--can be converted from green to red following exposure to blue-green light (450-500 nm). This photoconversion occurs in both insect and mammalian cells and is enhanced in a low oxygen environment. The red fluorescent GCaMPs retained calcium responsiveness, albeit with reduced sensitivity. We identified several amino acid residues in GCaMP important for photoconversion and generated a GCaMP variant with increased photoconversion efficiency in cell culture. This light-induced spectral shift allows the ready labeling of specific, targeted sets of GCaMP-expressing cells for functional imaging in the red channel. Together, these findings indicate the potential for greater utility of existing GCaMP reagents, including transgenic animals.

Publication types

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

MeSH terms

  • Animals
  • Animals, Genetically Modified
  • Calcium / metabolism
  • Calmodulin / chemistry*
  • Calmodulin / genetics
  • Calmodulin / metabolism
  • Color
  • Drosophila melanogaster
  • Fluorescence
  • Green Fluorescent Proteins / chemistry*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HEK293 Cells
  • Humans
  • Myosin-Light-Chain Kinase / chemistry*
  • Myosin-Light-Chain Kinase / genetics
  • Myosin-Light-Chain Kinase / metabolism
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Photochemical Processes*
  • Recombinant Fusion Proteins / chemistry*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism

Substances

  • Calmodulin
  • Peptide Fragments
  • Recombinant Fusion Proteins
  • Green Fluorescent Proteins
  • Myosin-Light-Chain Kinase
  • Calcium