MOLECULAR MECHANISMS OF RETINA SPECIFIC GENE EXPRESSION

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Differential expression of genes in a particular tissue, such as retina, is achieved by combinatoriaI action of transcription factors. Nrl is an evolutionarily-conserved bZIP transcription factor, identified in our laboratory by subtraction cloning. In the adult, high levels of Nrl transcripts are detected only in the retina. The Nrl protein shows strong homology to the product of a transforming oncogene, v-maf. The proteins of the Maf-Nrl subfamily recognize a long AP-1 like DNA sequence element, are shown to heterodimerize with selected bZIP proteins in vitro, and implicated in tissue-specific gene regulation. Several lines of evidence suggest that Nrl is involved in rhodopsin gene regulation. The underlying hypothesis is that in the adult retina Nrl plays a major role in regulating the expression of gene products that are needed for the appropriate fiinctioning of photoreceptors and other neurons. Since protein-protein interactions are a major determinant of transcriptional activity and can generate tremendous flexibility in target site selection, the objective of this proposal is to identify the proteins that specifically interact with Nrl and to elucidate the biological relevance of these interactions. To accomplish this, Specific Aim 1 proposes to evaluate whether any of the known bZIP proteins of the Fos, Jun, Maf-Nrl and NF-E2 p45 subfamilies is expressed in the adult retina and interacts with Nrl to modulate gene expression. Specific Aim 2 focuses on the purification and characterization of retinal proteins that bind to the Nrl-response element in the rhodopsin promoter. Specific Aim 3 employs a yeast two- hybrid approach with the "Nrl-bZIP bait" to identify the proteins that productively heterodimerize with Nrl and may be involved in regulating different sets of genes in the retina. The long-term objective of P.I.'s research is to understand the molecular events involved in the pathogenesis of eye diseases and eventually assist in the design of gene-based therapeutic strategies. Transcription factors have become attractive targets for mechanism- based design of drugs, which can be used to modulate specific biochemical pathways. In that direction, the proposed studies are designed to identify transcriptional regulatory proteins that together with Nrl mediate tissue- or cell type- specific gene expression in the retina, and should provide mechanistic insights into the molecular basis of congenital and inherited retinal disorders.