MOLECULAR CHARACTERIZATION OF THE RHODOPSIN GENE

视紫红质是视杆细胞外节的主要膜蛋白 感光细胞，启动复杂的级联事件相关 通过吸收光子来实现视觉的过程。 的 视紫红质基因的结构特征 以及调节其表达的分子机制对细胞生存的影响 感光细胞是一个重要的神经生物学问题， 相对而言，我们所知甚少。 在小鼠视网膜中， rds（视网膜变性缓慢）突变与a） 缺乏97%的正常量的视紫红质; B）视杆细胞的失效 外节发育;和c）遗传编程的早熟细胞 视杆细胞死亡。 杂合子（rds/+）只有一半 视紫红质水平正常，且形态上较短， 组织良好的杆外节。 总的来说，这些数据表明， RDS突变或者是与所述突变相邻的“顺式作用”调节基因， 视紫红质基因或它实际上是在视紫红质的结构基因内。 我们的总体目标是定义和表征视紫红质基因， 其在正常和RDS小鼠发育过程中的表达。 我们构建了一个 小鼠视网膜cDNA表达文库，克隆和部分测序小鼠 视紫红质cDNA，并获得了单克隆和单特异性抗体 抵抗视紫红质 我们现在建议：a）描述两者的特征 通过北方和西方的定性和定量分析， 视紫红质基因产物（mRNA和视蛋白）在正常发育过程中， B）克隆和测序来自rds小鼠的视紫红质基因， 正常和RDS小鼠的基因组文库;和c）确定染色体 使用Southern杂交法对小鼠视紫红质基因进行了定位和精确位点分析。 小鼠-中国仓鼠体细胞限制性内切酶片段分析 杂交染色体组和来自重组近交系小鼠。 的 还将监测其它视网膜特异性基因的表达， 为了表征RDS突变的生化表型， 感光细胞的发育和退化。 获得的结果 将对遗传性失明的研究具有重要意义 (e.g.视网膜色素变性），并将提供哺乳动物模型，用于研究 与遗传控制相关的分子事件 在中枢神经系统中， 系统

Rhodopsin, the major integral membrane protein of the rod outer segment of photoreceptor cells, initiates the complex cascade of events associated with the process of vision, by the absorption of a photon of light. The relationship of both the structural characteristics of the rhodopsin gene and the molecular mechanisms regulating its expression, to the survival of photoreceptor cells is an important neurobiological problem about which relatively little is known. In the mouse retina, the homozygous presence of the rds (retinal degeneration slow) mutation is associate with a) the absence of 97% of the normal amounts of rhodopsin; b) the failure of rod outer segments to develop; and c) the genetically programmed premature cell death of rod photoreceptor cells. The heterozygote (rds/+) has only half the level of rhodopsin as normal and has morphologically shorter and less well organized rod outer segments. Collectively, these data indicate that the rds mutation is either a "cis acting" regulatory gene adjacent to the rhodopsin gene or it is actually within the structural gene for rhodopsin. Our overall objective is to define and characterize the rhodopsin gene and its expression during development in normal and rds mice. We constructed a mouse retinal cDNA expression library, cloned and partially sequenced mouse rhodopsin cDNA and have obtained monoclonal and monospecific antibodies against rhodopsin. We are now proposing to: a) characterize both qualitatively and quantitatively by Northern and Western alalysis, the rhodopsin gene products (mRNA and opsin) during development of normal, heterozygote and rds mice; b) clone and sequence the rhodopsin gene from genomic libraries of normal and rds mice; and c) determine the chromosomal assignment and the exact locus of the mouse rhodopsin gene using Southern Analysis of restriction fragments from a mouse-Chinese hamster somatic cell hybrid chromosome panel and from recombinant inbred lines of mice. The expression of other retina specific genes will also be monitored in order to characterize the biochemical phenotype of the rds mutation during the development and degeneration of photoreceptor cells. The results obtained from this study will have importance for the study of hereditary blindness (e.g. retinitis pigmentosa) and will provide a mammalian model for studying the exact molecular events associated with genetically controlled degeneration of a unique population of neurons, in the central nervous system.