Of the two classes of vertebrate photoreceptor cells, cone photoreceptors are perhaps the most important with respect to vision since they function in bright light, subserve interpretation of color and are responsible for fine acuity. Relatively little is known, however, concerning the molecular foundations for cone photoreceptor identity and specificity. The primary goal of our continuing research is to define the molecular bases of structural, compositional and functional properties of cone photoreceptors and their interactions with the IPM using cell biological approaches. The emphasis of this proposal is placed on elucidating the molecular nature of """"""""cone matrix sheaths"""""""" (CMS), structurally distinct domains of chondroitin 6-sulfate-rich IPM that encapsulate cone outer segments. Although the precise function of CMSs has not yet been elucidated, studies from our laboratory suggest that they may be essential to cone outer segment viability and that maintenance of retinal adhesion may depend, in part, upon continuous synthesis and turnover of CMS- associated proteoglycans. In the studies proposed, we will utilize our ability to generate enriched preparations of cMSs to isolate, selectively purify ad characterize their soluble and insoluble constituents using biochemical techniques. Monoclonal and polyclonal antibodies will be generated against CMS- specific molecules and will be utilized subsequently to determine the molecular substructure of CMSs, to screen cDNA libraries, to assess changes in CMS composition following pharmacologic perturbation and to monitor their expression during retinal differentiation and their fate in various retinal pathologies. A porcine cone photoreceptor-enriched cDNA library will be generated and cone-specific clones will be identified and isolated. The incorporation of 35SO4 into CMS-associated molecules will be assessed by light microscopic autoradiography and SDS-PAGE fluorography to provide information pertaining to the molecular weights, metabolism and turnover of sulfated CMS-associated molecules. The effects of intravitreal injections of beta-D-xyloside (a compound that interferes with proteoglycan synthesis and secretion) on the incorporation of sulfate into CMS-associated molecules, on cone photoreceptor function and structure and on retinal adhesion will be determined morphologically and biochemically. CMS-specific molecules will be monitored immunocytochemically in developing human and porcine retinae in order to document the expression of specific molecular species and to define critical developmental periods when CMSs are established. In addition, the fate of CMS-specific molecules in humans and animals with hereditary and pathologic diseases, including retinitis pigmentosa and mucopolysaccharidosis, will be monitored immunocytochemically in order to define the relationship between the survival of cone photoreceptors and the integrity of CMSs. It is anticipated that successful completion of the above studies will add significantly to our overall understanding of the dynamic functions of cone photoreceptors and cone matrix sheaths in normal and diseased retinae.
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