We have established the visual system of the unicellular eukaryote, Clamydomonas reinhardtii, as a model for the study of vision. Its rhodopsin receptor appears to be homologous with that of bovine. We propose to use the ease of genetic, behavioral, and biochemical manipulation of Chlamydomonas to improve our knowledge of the molecular mechanisms of vision. The nature of the retinal environment in its opsin and the mechanism of transduction will be studied with a series of retinal analogs by behavior and in vivo differential spectroscopy. The receptor protein and its membrane will be isolated for related spectroscopic information. The components of Chlamydomonas' visual processing that shape, communicate and maintain its visual signal to its flagella are also likely to be homolgous to the similar components found in higher organism photoreceptors. As is true for other rhodoposin receptors the signal has steps in the form of ionic currents. Particularly suitable for cells of this size are the attached- and whole-cell clamp electrophysiological techniques. Several of the ionic pathway components of its visual signal processor will be identified by combining this approach with pharmacological, light, ionic and mutant modifications of conditions. An extensive search for mutants that effect any aspect of the visual process will be started. At the same time search for specific mutants of the opsin and the hypothesized retinal isomerase will be begun. The ease of obtaining mutants which enable isolation of each protein involved in the transduction process is a very attractive feature of this model system. The work proposed in this grant application should yield information relevant to the molecular details of transduction, signal processing, biosynthesis of chromophore, cell maintenance, effect of nutrition, influence of lipids, adapation, and cell homeostasis. Understanding these molecular mechanisms should make it possible to understand the effects of some genetic diseases, nutrition, light, and drugs on the transduction portion of the visual process.
