Tuning FlaSh: redesign of the dynamics, voltage range, and color of the genetically encoded optical sensor of membrane potential.
The optical voltage sensor FlaSh, made from a fusion of a GFP "reporter domain" and a voltage-gated Shaker K(+) channel "detector domain," has been mutagenically tuned in both the GFP reporter and channel detector domains. This has produced sensors with improved folding at 37 degrees C, enabling use in mammalian preparations, and yielded variants with distinct spectra, kinetics, and voltage dependence, thus expanding the types of electrical signals that can be detected. The optical readout of FlaSh has also been expanded from single wavelength fluorescence intensity changes to dual wavelength measurements based on both voltage-dependent spectral shifts and changes in FRET. Different versions of FlaSh can now be chosen to optimize the detection of either action potentials or synaptic potentials, to follow high versus low rates of activity, and to best reflect electrical activity in cell types with distinct voltages of operation.
Guerrero
Giovanna
G
Department of Molecular and Cell Biology, University of California-Berkeley, Berkeley, CA 94720-3200, USA.
Siegel
Micah S
MS
Roska
Botond
B
Loots
Eli
E
Isacoff
Ehud Y
EY
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology*; D008856:Microscopy, Fluorescence / Q000379:methods; D016297:Mutagenesis, Site-Directed; D009474:Neurons / Q000502:physiology; D055095:Optics and Photonics; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism; D024642:Potassium Channels, Voltage-Gated; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000502:physiology*; D051637:Shaker Superfamily of Potassium Channels
D008164:Luminescent Proteins; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011993:Recombinant Fusion Proteins; D051637:Shaker Superfamily of Potassium Channels; D049452:Green Fluorescent Proteins