MOLECULAR MECHANISMS OF RETINA SPECIFIC GENE EXPRESSION

基因在特定组织中的差异表达，例如 视网膜，是通过转录因子的组合定向作用实现的。 Nrl是进化上保守的bZIP转录因子， 在我们的实验室通过消减克隆鉴定。在成年人中，高 仅在视网膜中检测到Nr 1转录物的水平。全国劳资关系委员会 蛋白质显示出与转化产物的强同源性， 癌基因v-maf。Maf-Nrl亚家族的蛋白质识别长的 AP-1样DNA序列元件，显示与 选择bZIP蛋白在体外，并牵连在组织特异性基因 调控几条证据表明，Nrl参与了 视紫红质基因调控 潜在的假设是， 成人视网膜Nrl在调节 基因产物，需要适当的fintioning 光感受器和其他神经元。 由于蛋白质-蛋白质相互作用 是转录活性的主要决定因素， 在目标地点的选择上具有巨大的灵活性， 一个新的建议是确定蛋白质的具体相互作用， 并阐明这些相互作用的生物学意义。 为了实现这一目标，具体目标1建议评估是否有任何 已知的Fos、Jun、Maf-Nrl和NF-E2 p45的bZIP蛋白 亚家族在成人视网膜中表达，并与Nrl相互作用， 调节基因表达。具体目标2侧重于纯化 以及与Nr 1反应结合的视网膜蛋白的表征 视紫红质启动子中的元素。具体目标3采用酵母双- 使用“Nrl-bZIP诱饵”的混合方法来鉴定 与Nrl产生异二聚体，并可能参与调节 视网膜中不同的基因组 P.I.的长期目标。的研究是为了了解 参与眼病发病机制的分子事件， 最终帮助设计基于基因的治疗策略。 转录因子已成为有吸引力的目标机制- 基于药物设计，可用于调节特定的 生化途径在这方面，拟议的研究是 旨在识别转录调节蛋白， 与Nrl介导的组织或细胞类型特异性基因表达， 视网膜，并应提供机制的见解，分子 先天性和遗传性视网膜疾病的基础。

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.