During eye development, specific cell types differentiate from progenitor cells of diverse embryonic origin, and cell-cell interactions orchestrate the formation of a highly efficient, working sensory organ, in ways which are not understood. Later, the ocular cell types must maintain their differentiated phenotype for proper health and operation of the eye. Several diseases leading to loss of sight result from differentiation changes, for example, macular degeneration in the retina, cataracts of the lens and transdifferentiation of the retinal pigment epithelium (proliferative vitreoretinal disorders). Cadherins are calcium dependent cell-cell adhesion molecules that are essential for mammalian development. l have recently shown that cadherin cell-cell adhesion molecules control the differentiation state in retinal pigment epithelial cells; ectopic expression of E-cadherin in a rat retinal pigment epithelium cell line, RPE-J, results in a phenotypic transformation that includes reorganization of constitutive proteins and induced synthesis of specific mRNAS and proteins producing basal-lateral Na+, K+-ATPase polarity, ankyrin isoform switching, desmosome junctional complex assembly and keratin filament assembly. Using these observations, we now propose to analyze mechanisms for determining the differentiated state of the retinal pigment epithelium and extend these observations to other cell types of the eye. Our specific goals in the proposed studies are to: (i) identify cadherin sequences required for retinal pigment epithelium differentiation. Chimeric cadherins will be generated between endogenous retinal pigment epithelium cadherin and E-cadherin and expressed by transfection in RPE-J cells to define the sequences required for inducing phenotypic changes. (ii) Identify cadherin-associated molecules which may transduce cadherin- induced differentiation cues of the retinal pigment epithelium. After defining the sequences which are utilized during the cadherin-induced differentia~on program, biochemical and yeast two hybrid methods will be used to identify proteins which interact with these sequences. (iii) Analyze cadherin-induced differentiation the lens epithelium. Like retinal pigment epithelial cells, lens epithelial cells do not endogenously express E-cadherin. We will test the generality of the cadherin-induced differentiation program in this ocular cell type. (iv) Identify novel cadherins which may establish and maintain the differentiated state of lens epithelial cells and retinal cell types. We have initiated a search for novel cadherins which are expressed in ocular tissues. Preliminary evidence from my laboratory shows that there are cadherin molecules expressed in the retina which have not been previously described there. We propose an extensive screen for novel cadherins using RT-PCR methods, and to clone full length cDNAs encoding these novel cadherins. The results of these studies will provide a detailed mechanistic framework of cadherin- induced differentiation changes during eye morphogenesis and will identify potential sites in the differentiation program which fail during pathophysiological processes in eye tissues.
