Channelrhodopsin-1 Initiates Phototaxis and Photophobic Responses in Chlamydomonas by Immediate Light-Induced Depolarization[W]

Channelrhodopsins (CHR1 and CHR2) are light-gated ion channels acting as sensory photoreceptors in Chlamydomonas reinhardtii. In neuroscience, they are used to trigger action potentials by light in neuronal cells, tissues, or living animals. Here, we demonstrate that Chlamydomonas cells with low CHR2 content exhibit photophobic and phototactic responses that strictly depend on the availability of CHR1. Since CHR1 was described as a H+-channel, the ion specificity of CHR1 was reinvestigated in Xenopus laevis oocytes. Our experiments show that, in addition to H+, CHR1 also conducts Na+, K+, and Ca2+. The kinetic selectivity analysis demonstrates that H+ selectivity is not due to specific translocation but due to selective ion binding. Purified recombinant CHR1 consists of two isoforms with different absorption maxima, CHR1505 and CHR1463, that are in pH-dependent equilibrium. Thus, CHR1 is a photochromic and protochromic sensory photoreceptor that functions as a light-activated cation channel mediating phototactic and photophobic responses via depolarizing currents in a wide range of ionic conditions.

[1]  Peter Hegemann,et al.  Algal sensory photoreceptors. , 2008, Annual review of plant biology.

[2]  H. Fukuzawa,et al.  Archaeal-type rhodopsins in Chlamydomonas: model structure and intracellular localization. , 2003, Biochemical and biophysical research communications.

[3]  J. Bausch,et al.  Monoclonal antibodies. , 1990, Bioprocess technology.

[4]  R. Molday,et al.  Monoclonal antibodies to rhodopsin: characterization, cross-reactivity, and application as structural probes. , 1983, Biochemistry.

[5]  K. Kindle High-frequency nuclear transformation of Chlamydomonas reinhardtii. , 1990, Proceedings of the National Academy of Sciences of the United States of America.

[6]  P. Hegemann,et al.  Light‐induced stop response in Chlamydomonas reinhardtii: Occurrence and adaptation phenomena , 1989 .

[7]  P. Hegemann,et al.  REVERSIBLE BLEACHING OF Chlamydomonas reinhardtii RHODOPSIN in vivo , 1988, Photochemistry and photobiology.

[8]  P. Hegemann,et al.  In vitro identification of rhodopsin in the green alga Chlamydomonas. , 1991, Biochemistry.

[9]  U. Hansen,et al.  Reaction kinetic parameters for ion transport from steady-state current-voltage curves. , 1987, Biophysical journal.

[10]  Carol Sanger,et al.  Editing , 2020, Every Day I Write the Book.

[11]  Volker Wagner,et al.  Proteomic Analysis of the Eyespot of Chlamydomonas reinhardtii Provides Novel Insights into Its Components and Tactic Movements[W] , 2006, The Plant Cell Online.

[12]  P. Hegemann,et al.  The nature of rhodopsin-triggered photocurrents in Chlamydomonas. II. Influence of monovalent ions. , 1996, Biophysical Journal.

[13]  Oliver P. Ernst,et al.  Photoactivation of Channelrhodopsin* , 2008, Journal of Biological Chemistry.

[14]  J. Spudich The multitalented microbial sensory rhodopsins. , 2006, Trends in microbiology.

[15]  D. Sanders,et al.  Calcium-potassium selectivity: kinetic analysis of current-voltage relationships of the open, slowly activating channel in the vacuolar membrane of Vicia faba guard-cells , 1998, Planta.

[16]  Krzysztof Palczewski,et al.  Role of the conserved NPxxY(x)5,6F motif in the rhodopsin ground state and during activation , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[17]  Jureepan Saranak,et al.  A rhodopsin is the functional photoreceptor for phototaxis in the unicellular eukaryote Chlamydomonas , 1984, Nature.

[18]  Peter Berthold,et al.  An engineered Streptomyces hygroscopicus aph 7" gene mediates dominant resistance against hygromycin B in Chlamydomonas reinhardtii. , 2002, Protist.

[19]  A. Glass,et al.  Potassium Fluxes in Chlamydomonas reinhardtii (I.Kinetics and Electrical Potentials) , 1995, Plant physiology.

[20]  K. Jung The Distinct Signaling Mechanisms of Microbial Sensory Rhodopsins in Archaea, Eubacteria and Eukarya † , 2007, Photochemistry and photobiology.

[21]  Two light-transducing membrane proteins: bacteriorhodopsin and the mammalian rhodopsin. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[22]  P. Hegemann,et al.  Control of phobic behavioral responses by rhodopsin-induced photocurrents in Chlamydomonas. , 1997, Biophysical journal.

[23]  J. Ruppersberg Ion Channels in Excitable Membranes , 1996 .

[24]  T. Ishizuka,et al.  Kinetic evaluation of photosensitivity in genetically engineered neurons expressing green algae light-gated channels , 2006, Neuroscience Research.

[25]  R. Kamiya,et al.  The sensitivity of chlamydomonas photoreceptor is optimized for the frequency of cell body rotation. , 2001, Plant & cell physiology.

[26]  R. Uhl,et al.  How Chlamydomonas keeps track of the light once it has reached the right phototactic orientation. , 1997, Biophysical journal.

[27]  A. Dizhoor,et al.  Ectopic Expression of a Microbial-Type Rhodopsin Restores Visual Responses in Mice with Photoreceptor Degeneration , 2006, Neuron.

[28]  J. Spudich,et al.  Sensory Rhodopsin Signaling in Green Flagellate Algae , 2005 .

[29]  Larry Simpson,et al.  Mitochondrial proteins and complexes in Leishmania and Trypanosoma involved in U-insertion/deletion RNA editing. , 2004, RNA.

[30]  T. Oertner,et al.  Optical induction of synaptic plasticity using a light-sensitive channel , 2007, Nature Methods.

[31]  K. Deisseroth,et al.  Millisecond-timescale, genetically targeted optical control of neural activity , 2005, Nature Neuroscience.

[32]  P. Hegemann,et al.  Probing visual transduction in a plant cell: Optical recording of rhodopsin-induced structural changes from Chlamydomonas reinhardtii. , 1990, Biophysical journal.

[33]  E. Govorunova,et al.  Photoinduced electric currents in carotenoid-deficient Chlamydomonas mutants reconstituted with retinal and its analogs. , 1994, Biophysical journal.

[34]  G. Throm,et al.  Phototaktische Untersuchungen an Chlamydomonas reinhardii Dangeard in homokontinuierlicher Kultur , 1971, Archiv für Mikrobiologie.

[35]  Peter Hegemann,et al.  "Vision" in single-celled algae. , 2004, News in physiological sciences : an international journal of physiology produced jointly by the International Union of Physiological Sciences and the American Physiological Society.

[36]  P. Hegemann,et al.  Evidence for a light-induced H(+) conductance in the eye of the green alga Chlamydomonas reinhardtii. , 2002, Biophysical journal.

[37]  E. Bamberg,et al.  Channelrhodopsin-2, a directly light-gated cation-selective membrane channel , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[38]  E. Bamberg,et al.  Light Activation of Channelrhodopsin-2 in Excitable Cells of Caenorhabditis elegans Triggers Rapid Behavioral Responses , 2005, Current Biology.

[39]  H. Chiel,et al.  Fast noninvasive activation and inhibition of neural and network activity by vertebrate rhodopsin and green algae channelrhodopsin. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[40]  R. Mahendran,et al.  RNA editing by cytidine insertion in mitochondria of Physarum polycephalum , 1991, Nature.

[41]  J. Rosenbaum,et al.  Polarity of flagellar assembly in Chlamydomonas , 1992, The Journal of cell biology.

[42]  H. Khorana,et al.  A single amino acid substitution in rhodopsin (lysine 248----leucine) prevents activation of transducin. , 1988, The Journal of biological chemistry.

[43]  P. Hegemann,et al.  The nature of rhodopsin-triggered photocurrents in Chlamydomonas. I. Kinetics and influence of divalent ions. , 1996, Biophysical journal.

[44]  K. Palczewski,et al.  Signaling States of Rhodopsin , 2003, The Journal of Biological Chemistry.

[45]  Hartmann Harz,et al.  Rhodopsin-regulated calcium currents in Chlamydomonas , 1991, Nature.

[46]  Ernst Bamberg,et al.  Spectral characteristics of the photocycle of channelrhodopsin-2 and its implication for channel function. , 2008, Journal of molecular biology.

[47]  G. Nagel,et al.  Light-Induced Activation of Distinct Modulatory Neurons Triggers Appetitive or Aversive Learning in Drosophila Larvae , 2006, Current Biology.

[48]  Feng Zhang,et al.  Channelrhodopsin-2 and optical control of excitable cells , 2006, Nature Methods.

[49]  O. Sineshchekov,et al.  Photoreceptor electric potential in the phototaxis of the alga Haematococcus pluvialis , 1978, Nature.

[50]  E. Bamberg,et al.  Channelrhodopsin-1: A Light-Gated Proton Channel in Green Algae , 2002, Science.

[51]  Oleg A. Sineshchekov,et al.  Two rhodopsins mediate phototaxis to low- and high-intensity light in Chlamydomonas reinhardtii , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[52]  J. Spudich,et al.  Microbial rhodopsins: functional versatility and genetic mobility. , 2006, Trends in microbiology.

[53]  O. Andersen Kinetics of ion movement mediated by carriers and channels. , 1989, Methods in enzymology.

[54]  M. Schroda,et al.  The HSP70A promoter as a tool for the improved expression of transgenes in Chlamydomonas. , 2000, The Plant journal : for cell and molecular biology.

[55]  Kwang-Hwan Jung,et al.  Chlamydomonas sensory rhodopsins A and B: cellular content and role in photophobic responses. , 2004, Biophysical journal.

[56]  Ier Halldal Action Spectra of Phototaxis and Relation Problem in Volvocalcs, Ulva‐Gamete and Dinophyceae , 1958 .