| Research Abstract |
Many vertebrates have several types of photoreceptive cells ; rods and cones in the retina, and pinealocytes in the pineal gland. It is important to understand the difference in molecular mechanism responsible for the diverged cellular functions ; that is, rods for scotopic vision, cones for color vision, and pinealocytes for circadian regulation of biological clock. This project aimed at comparative studies on the photon-signal transduction processes in the three types of photoreceptive cells, particularly at the structures and functions of photoreceptive pigments and signal-transducing G-proteins in the cells.
[Rod] To identify the physiological roles of lipid modifications of transducin alpha-and betagamma-subunits (Talpha and Tbetagamma), we studied the mechanism of the subunit interaction and the membrane association. It ws clearly shown that the isoprenyl and methyl groups of Tgamma regulate the efficiency of photon-signal transduction by affecting 1) Talpha-Tbetagamma coupling th
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rough direct contact with alpha-subunit, and 2) the interaction between transducin and cellular membranes. Monoclonal antibodies recognizing the structure of the N-terminal fatty acid of Talpha stained almost all of the rod outer segments in a bovine retina, suggesting that a single rod cell contains the four isoforms of Talpha modified with four kinds of fatty acids. The target of recoverin was shown to be rhodopsin kinase. The complex of calcium-recoverin-rhodopsin kinase was stabilized by the membrane association, by which the kinase activity was effectively inhibited.
[Cone] Comparative studies on cone and rod photoreceptive pigments by low temperature spectroscopy, laser flash photolysis, and circular dichroism showed that the life times of the active intermediates of cone pigments are much faster than that of rhodopsin. This clearly indicates that the molecular property of cone pigment is one of the major determinants for physiological photoresponse of cones. Photochemical reactions of chimeric mutants composed of rod-and cone-type pigments demonstrated that the structures of helices 1 through 3 determines the life time of the active intermediate of photoreceptive pigments.
[Pinealocytes] Little is known about the photon-signal transduction process in a photosensitive pineal gland. We found opsin-like immunoreaction in the gland by immunohistochemistry. Then we isolated the cDNA encoding a pineal specific opsin-like protein. Reconstitution experiment with 11-cis-retinal revealed its blue-light sensitivity, and thus we named it "pinopsin" after pineal opsin. This is the first demonstration of an opsin expressed in non-retinal tissues. Less
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