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周期的基因 视觉反应。已知视觉级联的几个成员是 被光激活后被磷酸化。以此类推 系统，这种磷酸化被认为是负责一个 很大一部分是对响应的调制。因此，我们计划 专注于分离编码调节性激酶的基因。 导致视觉级联过度激活的突变会导致 视网膜变性。因此，对消费者反应水平的监管 视觉系统不仅对正常的视力至关重要，而且对维持视力也至关重要 光感受器细胞的活性。影响人类免疫功能水平的突变 视紫红质的磷酸化也被认为会导致退化。 因此，我们正在寻找的丝氨酸/苏氨酸激酶可能是关键 视网膜退行性疾病的玩家。 为了分离丝氨酸/苏氨酸蛋白激酶基因，我们设计了 编码保守氨基酸结构域的两个简并寡核苷酸 VI和VIII存在于激酶家族的所有成员中。聚合酶链式反应将 对眼睛特异序列富含多倍的cDNAs 利用突变蝇eya的Head mRNA进行消减杂交 不会发育成眼睛。扩增的序列将被分离和筛选 通过测序获得了激活酶。任何发现的激酶都将被进一步表征 以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.