ISOLATION OF REGULATORY KINASES FROM THE VISUAL SYSTEM OF DROSOPHILA

我们感兴趣的是分离和表征调节细胞凋亡的基因。 视觉反应 视觉级联的几个成员是已知的 磷酸化响应于光的激活。 通过类推 系统，据认为，这种磷酸化是负责 很大一部分的反应调制。 因此，我们计划 集中于分离编码调节激酶的基因。 导致视觉级联反应过度激活的突变会导致 视网膜变性 因此，调节反应的水平， 视觉系统不仅对正常视力至关重要， 光感受器细胞的活力。 突变影响的水平 视紫红质的磷酸化也已知引起变性。 因此，我们正在寻找的丝氨酸/苏氨酸激酶可能是关键 视网膜变性疾病的参与者。 为了分离丝氨酸/苏氨酸蛋白激酶基因，我们设计了 两个编码保守氨基酸结构域的简并寡核苷酸 VI和VIII在激酶家族的所有成员中发现。 PCR反应将 在cDNA上进行，所述cDNA通过多重PCR富集眼睛特异性序列。 使用来自突变果蝇Eya的头部mRNA进行消减杂交， 不会长眼睛。 扩增序列将被分离和筛选 通过测序来寻找激酶。 任何发现的激酶将进一步表征 来评估它们在调节视觉系统中的作用。

We are interested in isolating and characterizing genes which modulate the visual response. Several members of the visual cascade are known to be phosphorylated in response to activation by light. By analogy to other systems, it is thought that this phosphorylation is responsible for a large part of the modulation of the response. We therefore plan to concentrate on the isolation of genes which encode regulatory kinases. Mutations which cause excessive activation of the visual cascade lead to retinal degeneration. Thus, regulation of the level of response of the visual system is critical not only for proper vision but also to maintain viability of the photoreceptor cells. Mutations which affect the level of phosphorylation of rhodopsin are also know to cause degeneration. Therefore, the serine/threonine kinases we are searching for may be key players in retinal degenerative diseases. In order to isolate serine/threonine protein kinase genes, we have designed two degenerate oligonucleotides which encode conserved amino acid domains VI and VIII found in all members of the kinase family. PCR reactions will be done on cDNA which is enriched for eye-specific sequences by multiple subtractive hybridizations using head mRNA from the mutant fly eya which does not develop eyes. Amplified sequences will be isolated and screened for kinases by sequencing. Any kinases found will be further characterized to assess their role in regulating the visual system.