The long-term interests of this laboratory are the molecular mechanisms underlying the development and function of rods and cones. The questions are how expression of photoreceptor genes is regulated and how these cell-specific proteins contribute to the delivery of rod and cone phototransduction responses. We will combine molecular and cell biology, confocal microscopy, electrophysiology and microspectrophotometry in a novel and powerful transgenic system, Xenopus, focusing on the visual pigments. We will investigate: (1) molecular mechanisms underlying photoreceptor noise; 2) visual pigment properties underlying the dynamics of rod and cone response; and (3) regulatory regions in opsin genes underlying rod- and cone-specific expression. The rapid and inexpensive production of transgenic Xenopus, coupled with the large size of photoreceptors suitable for suction electrode recording, cone-rich retina and photoreceptor-specific promoters establish Xenopus as an ideal model for the proposed studies. The ability to introduce opsin mutants that have altered stability or photochemistry into rod cells will allow us to address several longstanding issues about rod photoreceptor behavior: mechanisms controlling sensitivity, noise and response recovery in a comprehensive way. Moreover, the characterization of promoters for opsins in cones opens the way to study cone physiology and signal transduction in an experimentally tractable organism. One of the principal determinants of visual system performance is photoreceptor sensitivity, which is limited by the inherent noise of the photoreceptor. In fact, noise plays an important role in dark adaptation and pathophysiological conditions such as night blindness and retinal degeneration. Given the importance of noise in determining visual sensitivity and the wide range of opsin mutants that impair vision, we intend to study the properties of rhodopsin that determine the lifetime of activated rhodopsin (equivalent to Meta II) which plays a crucial role in terminating the cell's response kinetics and amplification, in order to further understand the factors that limit daylight vision.
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