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First published online 18 January 2005
doi: 10.1242/jcs.01633

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Phototactic activity in Chlamydomonas 'non-phototactic' mutants deficient in Ca²⁺-dependent control of flagellar dominance or in inner-arm dynein

¹ Department of Biological Sciences, Graduate School of Science, University of Tokyo, Hongo, Tokyo 113-0063, Japan
² Doctoral Course of Structural Biosciences, Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba 305-8572, Japan

View larger version (93K): [in a new window]   Fig. 1. Photoaccumulation in the wild type (left) and the lsp1 mutant (right). Cell suspensions are shown after being exposed to light incident from the top of the figure for 5 minutes. Wild-type cells accumulated toward the light, whereas lsp1 cells remained distributed uniformly.

View larger version (16K): [in a new window]   Fig. 2. Swimming directions of the wild-type cells and various phototaxis mutant cells 5 seconds after the phototactic light was turned on. (A-D) Polar histograms showing the swimming directions when the light (wild type, 8.8x1016 photons m-2 s-1; lsp1, ptx1 and ida1, 2.2x1018 photons m-2 s-1) was applied in the direction indicated by arrows. Bars show the proportions of cells swimming in the direction binned at 15°. The interval between dotted circles represents 5% of the cells. Data on a total of 420 cells were collected from three experiments. (E,F) Stimulus-response curves for the phototaxis in the wild type (squares) and three mutant cells (lsp1, circles; ptx1, triangles; ida1, crosses). The magnitudes of phototaxis are shown by photo-orientation index calculated from the direction of swimming relative to the direction of light. Dotted line shows the value when the cells exhibit no phototaxis. Each point represents the average and standard deviation from three experiments (140 cells in each experiment).

View larger version (16K): [in a new window]   Fig. 3. Photoreceptor current (PRC) in wild-type and lsp1 mutant cells. (A,B) Typical waveforms observed when a flash (3.0x1018 photons m-2) was applied to wild-type (A) and lsp1 (B) cells at the time indicated by arrows. (C) Light dependence of the PRC in wild type (squares) and lsp1 mutant (circles). Averages and standard deviations from four experiments are plotted.

View larger version (14K): [in a new window]   Fig. 4. Photophobic response of wild type (squares) and lsp1 mutant (circles). The proportions of the cells showing photophobic response was examined at the intensity of flash light indicated on the abscissa. Averages and standard deviations from four measurements each (18-94 cells in each experiment) are shown.

View larger version (27K): [in a new window]   Fig. 5. The effects of Ca2+ concentration on the dominance of the activity of cis and trans axonemes. (A,B) Superimposed frames from video records and associated interpretations of rotating wild-type cells. Cells tended to rotate with the eyespot pointing inwards in a solution containing 10-6 M Ca2+ (A) and outwards in Ca2+-free solution (B). Scale bar, 10 µm. (C) The proportion of reactivated cell models whose trans axoneme beats dominantly at various Ca2+ concentrations. Each point shows the average and standard deviation from six experiments on wild-type (squares), lsp1 (circles), ptx1 (triangles) and ida1 (crosses) cells. The number of cells counted in each experiment was 29-159.

View larger version (20K): [in a new window]   Fig. 6. Flagellar beat frequencies in demembranated and reactivated cell models. The histograms for wild-type (A), lsp1 (B) and ptx1 (C) cells are shown.

View larger version (29K): [in a new window]   Fig. 7. (A) A genomic DNA segment that rescues the lsp1 phenotype. Boxes represent the open reading frames predicted by the exon-prediction software Green Genie. The sites of restriction enzyme digestion by which rescue was abolished are indicated on the top. The triangle shows the site of the plasmid insertion. The lsp1 phenotype was not rescued when the first or the last exon of the predicted gene was deleted (bottom). (B) Predicted primary structure of Lsp1. Phosphorylation sites are indicated. The sequences have been deposited in DDBJ/EMBL/GenBank with accession number AD194902.

View larger version (26K): [in a new window]   Fig. 8. (A) Hypothetical trans and cis transducers that transmit information from the sensor to dynein. (B) Reciprocal activation of the cis and trans complex in positive (top) and negative (bottom) phototaxis.

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