Abstract
The function of the eyespot in phototaxis of the flagellate green alga Chlamydomonas reinhardtii Dangeard was studied using quantitative reflection confocal laser scanning microscopy and photoelectric measurements. The reflective properties of the eyespot and the photoreceptor current of the C. reinhardtii eyespot mutant ey 627, mt − were compared with those of Chlamydomonas strains possessing a well-developed eyespot. Under growth conditions in which strongly disorganized eyespots were observed in the mutant by electron microscopy, there was a significant reduction in the reflection intensity of the eyespot and in the amplitude ratio (500∶440 nm) of photoreceptor currents induced by flashes of 500- and 440-nm light in non-oriented cells. Photoelectrical responses of pre-oriented cells revealed that the latter effect is caused by an altered directional sensitivity of the antenna complex, whereas the functional state of the photoreceptor pigment is not strongly affected in mutant cells. Both the reflection intensity and the amplitude ratio of photoreceptor currents increased to the level of reference strains under conditions supporting the development of a well-organized eyespot in the mutant. Furthermore, incubation of the mutant with high concentrations of all-trans-retinal (10 μM), independent of whether carotenoid biosynthesis was inhibited or not, was found to increase the reflection intensity of the eyespot. An increase in the rate of photoorientation of the mutant occurred concomitant with the increase in the reflective properties of the mutant eyespot. These observations demonstrate the importance of an intact eyespot for interference reflection and absorption of phototactically active light, and thus for the directional sensitivity of the eyespot apparatus.
Similar content being viewed by others
Abbreviations
- HSM:
-
high-salt medium
Referencess
Beckmann, M., Hegemann, P. (1991) In vitro identification of rhodopsin in the green alga Chlamydomonas. Biochemistry 30, 3692–3697
Boscov, J.S., Feinleib, M.E. (1979) Phototactic response of Chlamydomonas to flashes of light. II. Response of individual cells. Photochem. Photobiol. 30, 499–505
Buder, J. (1917) Zur Kenntnis der phototaktischen Richtungsbewegung. Jahrb. Wiss. Bot. 58, 105–220
Derguini, F., Mazur, P., Nakanishi, K., Starace, D.M., Saranak, J., Foster, K.W. (1991) All-trans-retinal is the chromophore bound to the photoreceptor of the alga Chlamydomonas reinhardtii. Photochem Photobiol. 54, 1017–1021
Foster, K.W., Smyth, R.D. (1980) Light antennas in phototactic algae. Microbiol. Rev. 44, 572–630
Foster, K.W., Saranak, J., Patel, N., Zarilli, G., Okabe, M., Kline, T., Nakanishi, K. (1984) A rhodopsin is the functional photoreceptor for phototaxis in the unicellular eukaryote Chlamydomonas. Nature 311, 756–759
Foster, K.W., Saranak, J., Zarrilli, G. (1988) Autoregulation of rhodopsin synthesis in Chlamydomonas reinhardtii. Proc. Natl. Acad. Sci. USA 85, 6379–6383
Harz, H., Hegemann, P. (1991) Rhodopsin-regulated calcium currents in Chlamydomonas. Nature 351, 489–491
Hader, D.-P. (1979) Photomovement. In: Encyclopedia of plant physiology, N.S., vol. 7: Physiology of movements, pp. 268–309, Haupt, W., Feinleib, M.E., eds. Springer, Berlin Heidelberg New York
Hegemann, P., Hegemann, U., Foster, K.W. (1988) Reversible bleaching of Chlamydomonas reinhardtii rhodopsin in vivo. Photochem. Photobiol. 48, 123–128
Hegemann, P., Gärtner, W., Uhl, R. (1991) All-trans retinal constitutes the functional chromophore in Chlamydomonas rhodopsin. Biophys. J. 60, 1477–1489
Kreimer, G., Melkonian, M. (1990) Reflection confocal laser scanning microscopy of eyespots in flagellated green algae. Eur. J. Cell Biol. 53, 101–111
Kreimer, G., Brohsonn, U., Melkonian, M. (1991a) Isolation and partial characterization of the photoreceptive organelle for phototaxis of a flagellate green alga. Eur. J. Cell Biol. 55, 318–327
Kreimer, G., Kawai, H., Müller, D.G., Melkonian, M. (1991b) Reflective properties of the stigma in male gametes of Ectocarpus siliculosus (Phaeophyceae) studied by confocal laser scanning microscopy. J. Phycol. 27, 268–276
Kreimer, G., Marner, F.-J., Brohsonn, U., Melkonian, M. (1991c) Identification of 11-cis and all-trans-retinal in the photoreceptive organelle of a flagellate green alga. FEBS-Lett. 293, 49–52
Lien, T., Knutsen, G. (1976) Synchronized cultures of a cell wallless mutant of Chlamydomonas reinhardtii. Arch. Microbiol. 108, 189–194
Lien, T., Knutsen, G. (1979) Synchronous growth of Chlamydomonas reinhardtii (Chlorophyceae): a review of optimal conditions. J. Phycol. 15, 191–200
Litvin, F.F., Sineshchekov, O.A., Sineshchekov, V.A. (1978) Photoreceptor electric potential in the phototaxis of the alga Haematococcus pluvialis. Nature 271, 476–478
Mast, S.O. (1927) Structure and function of the eye-spot in unicellular and colonial organisms. Arch. Protistenkd. 60, 197–220
McFadden, G.I., Melkonian, M. (1986) Use of Hepes buffer for microalgal culture media and fixation for electron microscopy. Phycologia 25, 551–557
Melkonian, M. (1975) The fine structure of zoospores of Fritschiella tuberosa lyeng (Chaetophorineae, Chlorophyceae) with special reference to the flagellar apparatus. Protoplasma 86, 391–404
Melkonian, M. (1982) Effects of divalent cations on flagellar scales in the green flagellate Tetraselmis cordiformis. Protoplasma 111, 221–233
Melkonian, M., Robenek, H. (1980) Eyespot membranes of Chlamydomonas reinhardtii: a freeze-fracture study. J. Ultrastruct. Res. 72, 90–102
Melkonian, M., Robenek, H. (1984) The eyespot apparatus of flagellated green algae: a critical review. Prog. Phycol. Res. 3, 193–268
Morel-Laurens, N.M.L., Bird, D.J. (1984) Effects of cell division on the stigma of wild-type and an “eyeless” mutant of Chlamydomonas. J. Ultrastruc. Res. 87, 46–61
Morel-Laurens, N.M.L., Feinleib, M.E. (1983) Photomovement in an “eyeless” mutant of Chlamydomonas. Photochem. Photobiol. 37, 189–194
Nultsch, W., Hader, D.-P. (1988) Photomovement in motile microorganisms-II. Photochem. Photobiol. 47, 837–869
Nultsch, W., Throm, G., von Rimscha, I. (1971) Phototaktische Untersuchungen an Chlamydomonas reinhardtii Dangeard in homokontinuierlicher Kultur. Arch. Mikrobiol. 80, 351–369
Pringsheim, E.G. (1963) Farblose Algen. Gustav Fischer Verlag, Stuttgart
Reize, I.B., Melkonian, M. (1989) A new way to investigate living flagellated/ciliated cells in the light microscope: immobilization of cells in agarose. Bot. Acta 102, 145–151
Rüffer, U., Nultsch, W. (1991) Flagellar photoresponses of Chlamydomonas cells held on micropipettes: IL Change in flagellar beat pattern. Cell Motil. Cytoskeleton 18, 269–278
Sineshchekov, O.A. (1991a) Photoreception in unicellular flagellates: bioelectric phenomena in phototaxis. In: Light in biology and medicine, Proceedings of the III Conference of the European Society for Photobiology Budapest 1989, vol. 2, pp. 523–532, Douglas, R.H., Moan, J., Ronto, G., eds. Plenum Press, New York
Sineshchekov, O.A. (1991b) Electrophysiology of photomovements in flagellated algae. In: Biophysics of photoreceptors and photomovements, pp. 191–202, Lenci, F. ed. Plenum Press, New York
Sineshchekov, O.A., Govorunova, E.G., Litvin, F.F. (1989) Role of photosynthetic apparatus and stigma in the formation of spectral sensitivity of phototaxis in flagellated green algae. In Russ. Eng. abstr. Biofizika 34, 255–258
Sineshchekov, O.A., Litvin, F.F., Keszthelyi, L. (1990) Two components of photoreceptor potential in phototaxis of the flagellated green alga Haematococcus pluvialis. Biophys. J. 57, 33–39
Sineshchekov, O.A., Govorunova, E.G., Der, A., Keszthelyi, L., Nultsch, W. (1992) Photoelectric responses in phototactic flagellated algae measured in cell suspensions. J. Photochem. Photobiol. 13, 119–134
Sueoka, N., Chiang, K.S., Kates, J.R. (1967) Deoxyribonucleic acid replication in meiosis of Chlamydomonas reinhardtii. I. Isotopic transfer experiments with a strain producing eight zoospores. J. Mol. Biol. 25, 47–66
Surzycki, S. (1971) Synchronous grown cultures of Chlamydomonas reinhardtii. Methods Enzymol. 23, 67–73
Takahashi, T., Yoshihara, K., Watanabe, M., Kubota, M., Johnson, R., Derguini, F., Nakanishi, K. (1991) Photoisomerization of retinal at 13-ene is important for phototaxis of Chlamydomonas reinhardtii: simultaneous measurements of phototactic and photophobic responses. Biochem. Biophys. Res. Commun. 178, 1273–1279
Author information
Authors and Affiliations
Additional information
This study was supported by the Deutsche Forschungsgemeinschaft. O. A. Sineshchekov was supported by a Research Fellowship from the Alexander von Humboldt Foundation. The authors wish to thank U. Powalowski (Botanisches Institut, Universität zu Köln) for help with electron microscopy.
Rights and permissions
About this article
Cite this article
Kreimer, G., Overländer, C., Sineshchekov, O.A. et al. Functional analysis of the eyespot in Chlamydomonas reinhardtii mutant ey 627, mt − . Planta 188, 513–521 (1992). https://doi.org/10.1007/BF00197043
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00197043