Genetically encoded fluorescent voltage sensors using the voltage-sensing domain of Nematostella and Danio phosphatases exhibit fast kinetics

A substantial increase in the speed of the optical response of genetically encoded fluorescent protein voltage sensors (FP voltage sensors) was achieved by using the voltage-sensing phosphatase genes of Nematostella vectensis and Danio rerio. A potential N. vectensis voltage-sensing phosphatase was identified in silico. The voltage-sensing domain (S1-S4) of the N. vectensis homolog was used to create an FP voltage sensor called Nema. By replacing the phosphatase with a cerulean/citrine FRET pair, a new FP voltage sensor was synthesized with fast off kinetics (Tau(off)<5ms). However, the signal was small (ΔF/F=0.4%/200mV). FP voltage sensors using the D. rerio voltage-sensing phosphatase homolog, designated Zahra and Zahra 2, exhibited fast on and off kinetics within 2ms of the time constants observed with the organic voltage-sensitive dye, di4-ANEPPS. Mutagenesis of the S4 region of the Danio FP voltage sensor shifted the voltage dependence to more negative potentials but did not noticeably affect the kinetics of the optical signal.

[1]  Rodrigo Lopez,et al.  Clustal W and Clustal X version 2.0 , 2007, Bioinform..

[2]  E. K. Kosmidis,et al.  Three fluorescent protein voltage sensors exhibit low plasma membrane expression in mammalian cells , 2007, Journal of Neuroscience Methods.

[3]  Walther Akemann,et al.  Engineering of a Genetically Encodable Fluorescent Voltage Sensor Exploiting Fast Ci-VSP Voltage-Sensing Movements , 2008, PloS one.

[4]  M. Ohkura,et al.  A high signal-to-noise Ca2+ probe composed of a single green fluorescent protein , 2001, Nature Biotechnology.

[5]  Amiram Grinvald,et al.  VSDI: a new era in functional imaging of cortical dynamics , 2004, Nature Reviews Neuroscience.

[6]  Nicholas H. Putnam,et al.  Sea Anemone Genome Reveals Ancestral Eumetazoan Gene Repertoire and Genomic Organization , 2007, Science.

[7]  Botond Roska,et al.  Tuning FlaSh: redesign of the dynamics, voltage range, and color of the genetically encoded optical sensor of membrane potential. , 2002, Biophysical journal.

[8]  B M Salzberg,et al.  A large change in axon fluorescence that provides a promising method for measuring membrane potential. , 1973, Nature: New biology.

[9]  Hugh B. Nicholas,et al.  GeneDoc: a tool for editing and annotating multiple sequence alignments , 1997 .

[10]  Walther Akemann,et al.  Engineering and Characterization of an Enhanced Fluorescent Protein Voltage Sensor , 2007, PLoS ONE.

[11]  F. Jöbsis,et al.  Spectrophotometric studies on the pH of frog skeletal muscle. PH change during and after contractile activity , 1976, The Journal of general physiology.

[12]  S. Karlin,et al.  Prediction of complete gene structures in human genomic DNA. , 1997, Journal of molecular biology.

[13]  Yasushi Okamura,et al.  Improving membrane voltage measurements using FRET with new fluorescent proteins , 2008, Nature Methods.

[14]  Walther Akemann,et al.  Frontiers in Molecular Neuroscience Molecular Neuroscience Review Article Second and Third Generation Voltage-sensitive Fl Uorescent Proteins for Monitoring Membrane Potential , 2022 .

[15]  Sreekanth H. Chalasani,et al.  Imaging neural activity in worms, flies and mice with improved GCaMP calcium indicators , 2009, Nature Methods.

[16]  Walther Akemann,et al.  Engineering and Characterization of an Enhanced Fluorescent Protein Voltage Sensor , 2007, Neuroscience Research.

[17]  Leon Lagnado,et al.  A genetically-encoded reporter of synaptic activity in vivo , 2009, Nature Methods.

[18]  Javier Díez-García,et al.  Optical probing of neuronal circuit dynamics: genetically encoded versus classical fluorescent sensors , 2006, Trends in Neurosciences.

[19]  A. Kumánovics,et al.  Family ties of gated pores: evolution of the sensor module , 2002, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[20]  Yasushi Okamura,et al.  Phosphoinositide phosphatase activity coupled to an intrinsic voltage sensor , 2005, Nature.

[21]  Vincent A Pieribone,et al.  A genetically targetable fluorescent probe of channel gating with rapid kinetics. , 2002, Biophysical journal.

[22]  Thomas Knöpfel,et al.  Exploration of Fluorescent Protein Voltage Probes Based on Circularly Permuted Fluorescent Proteins , 2009, Front. Neuroeng..

[23]  W. N. Ross,et al.  Changes in Intracellular Free Calcium Concentration during Illumination of Invertebrate Photoreceptors , 1974, The Journal of general physiology.

[24]  F. Bezanilla,et al.  Detecting rearrangements of shaker and NaChBac in real-time with fluorescence spectroscopy in patch-clamped mammalian cells. , 2004, Biophysical journal.

[25]  Ehud Y Isacoff,et al.  A Genetically Encoded Optical Probe of Membrane Voltage , 1997, Neuron.

[26]  Walther Akemann,et al.  Optogenetic monitoring of membrane potentials , 2011, Experimental physiology.

[27]  Knut Holthoff,et al.  Rapid time course of action potentials in spines and remote dendrites of mouse visual cortex neurons , 2010, The Journal of physiology.

[28]  Yasushi Okamura,et al.  Enzyme Domain Affects the Movement of the Voltage Sensor in Ascidian and Zebrafish Voltage-sensing Phosphatases* , 2008, Journal of Biological Chemistry.

[29]  F Bezanilla,et al.  Charge-shift probes of membrane potential. Characterization of aminostyrylpyridinium dyes on the squid giant axon. , 1985, Biophysical journal.