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A genetically encoded Ca2+ indicator based on circularly permutated sea anemone red fluorescent protein eqFP578.
Shen, Yi; Dana, Hod; Abdelfattah, Ahmed S; Patel, Ronak; Shea, Jamien; Molina, Rosana S; Rawal, Bijal; Rancic, Vladimir; Chang, Yu-Fen; Wu, Lanshi; Chen, Yingche; Qian, Yong; Wiens, Matthew D; Hambleton, Nathan; Ballanyi, Klaus; Hughes, Thomas E; Drobizhev, Mikhail; Kim, Douglas S; Koyama, Minoru; Schreiter, Eric R; Campbell, Robert E.
Affiliation
  • Shen Y; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Dana H; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
  • Abdelfattah AS; Present address: Department of Neurosciences, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, OH, 4195, USA.
  • Patel R; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Shea J; Present address: Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
  • Molina RS; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
  • Rawal B; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
  • Rancic V; Department of Cell Biology and Neuroscience, Montana State University, Bozeman, MT, 59717, USA.
  • Chang YF; Department of Physiology, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada.
  • Wu L; Department of Physiology, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada.
  • Chen Y; LumiSTAR Biotechnology Incorporation, Nangang District, Taipei City, 115, Taiwan.
  • Qian Y; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Wiens MD; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Hambleton N; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Ballanyi K; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Hughes TE; Department of Chemistry, University of Alberta, Edmonton, Alberta, T6G 2G2, Canada.
  • Drobizhev M; Department of Physiology, University of Alberta, Edmonton, Alberta, T6G 2H7, Canada.
  • Kim DS; Department of Cell Biology and Neuroscience, Montana State University, Bozeman, MT, 59717, USA.
  • Koyama M; Department of Cell Biology and Neuroscience, Montana State University, Bozeman, MT, 59717, USA.
  • Schreiter ER; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
  • Campbell RE; Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, 20147, USA.
BMC Biol ; 16(1): 9, 2018 01 16.
Article in En | MEDLINE | ID: mdl-29338710
ABSTRACT

BACKGROUND:

Genetically encoded calcium ion (Ca2+) indicators (GECIs) are indispensable tools for measuring Ca2+ dynamics and neuronal activities in vitro and in vivo. Red fluorescent protein (RFP)-based GECIs have inherent advantages relative to green fluorescent protein-based GECIs due to the longer wavelength light used for excitation. Longer wavelength light is associated with decreased phototoxicity and deeper penetration through tissue. Red GECI can also enable multicolor visualization with blue- or cyan-excitable fluorophores.

RESULTS:

Here we report the development, structure, and validation of a new RFP-based GECI, K-GECO1, based on a circularly permutated RFP derived from the sea anemone Entacmaea quadricolor. We have characterized the performance of K-GECO1 in cultured HeLa cells, dissociated neurons, stem-cell-derived cardiomyocytes, organotypic brain slices, zebrafish spinal cord in vivo, and mouse brain in vivo.

CONCLUSION:

K-GECO1 is the archetype of a new lineage of GECIs based on the RFP eqFP578 scaffold. It offers high sensitivity and fast kinetics, similar or better than those of current state-of-the-art indicators, with diminished lysosomal accumulation and minimal blue-light photoactivation. Further refinements of the K-GECO1 lineage could lead to further improved variants with overall performance that exceeds that of the most highly optimized red GECIs.
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Full text: 1 Database: MEDLINE Main subject: Calcium / Luminescent Agents / Luminescent Proteins Limits: Animals / Humans Language: En Year: 2018 Type: Article
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PubMed Links
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Full text: 1 Database: MEDLINE Main subject: Calcium / Luminescent Agents / Luminescent Proteins Limits: Animals / Humans Language: En Year: 2018 Type: Article