Genetics of Endocytic Trafficking in the Drosophila Eye

描述(申请人提供)：细胞持续吸收营养和 信号分子。预定要降解的内化蛋白质被转送 从质膜到早期的内小体、多囊泡小体， 晚内体，最后到溶酶体。调控的分子机制 内吞转运中的这些步骤很重要，因为它们是由 许多遗传性疾病，包括切迪亚克-弗利加希综合征或赫尔曼斯基-普德拉克综合征。 果蝇复眼是一个很好的遗传分析模型系统 内吞贩卖的证据。这一过程所必需的许多基因的突变 可以被识别为眼睛颜色突变，因为它们还干扰 将生物合成的货物运送到颜料颗粒。老板的内部化 R7光感受器细胞的配体提供了跟踪内吞作用的直接检测方法 贩卖人口。这项提议旨在利用 果蝇的眼睛寻找内皮细胞运输的基因解剖 多细胞生物。 深橙色和(汽车)康乃馨眼睛颜色基因编码两个亚基。 在溶酶体交付和眼睛色素沉着的后期是必需的复合体。 在特定目标1中，该复合体的附加亚基在溶酶体中的比例 交付将以此为特征。在具体目标2中，具体缺陷在于 将确定DOR突变细胞的内吞和生物合成运输 用HRP融合标记不同隔区的超微结构水平 蛋白质。 利用溶酶体递送和眼睛颜色之间令人兴奋的联系 突变，特异性目标3提出了一个系统的突变筛查 眼睛颜色和内吞运输。除了额外的类似DOR的突变 在该途径中作用较晚的这一筛选也会产生干扰突变 在内吞转运的早期步骤中，例如，对MVB的调节 生物发生和功能。具体目标4提出了 DmVps28作为这种突变在内吞途径早期作用的一个例子。 这项工作的一个重要的长期目标是将不同的缺陷联系起来 将突变转移到改变内吞的遗传病的症状上 贩卖人口。为了直接比较果蝇模型系统中的表型， 特定的目标5是针对分离果蝇的突变 Hermansky-Pudlak综合征-1基因及其特征 内吞转运方面的缺陷。

DESCRIPTION (provided by applicant): Cells continuously take up nutrients and signaling molecules. Internalized proteins destined for degradation are routed from the plasma membrane through early endosomes, multivesicular bodies (MVBs), late endosomes and finally to lysosomes. The molecular mechanisms regulating these steps in endocytic trafficking are important, because they are altered by many genetic diseases including Chediak-Fligashi or Hermansky-Pudlak syndromes. The Drosophila compound eye is an excellent model system for a genetic analysis of endocytic trafficking. Mutations in many genes necessary for this process can be identified as eye color mutations because they also interfere with the delivery of biosynthetic cargo to pigment granules. Internalization of the Boss ligand into R7 photoreceptor cells provides a direct assay to follow endocytic trafficking. This proposal is aimed at exploiting these features of the Drosophila eye for a genetic dissection of endocytic trafficking in multicellular organisms. The (dor) deep orange and (car) carnation eye color genes encode two subunits of a complex that is necessary late in lysosomal delivery and eye pigmentation. In Specific Aim 1, the rote of additional subunits of this complex in lysosomal delivery will be characterized. In Specific Aim 2, the specific defects in endocytic and biosynthetic trafficking in dor mutant cells will be determined on the ultrastructural level by labeling different compartments with HRP fusion proteins. To exploit the exciting connection between lysosomal delivery and eye color mutations, Specific Aim 3 proposes a systematic screen for mutants affecting eye color and endocytic trafficking. Besides additional dor-like mutations acting late in the pathway, this screen will also yield mutations interfering with earlier steps in endocytic trafficking, for example, the regulation of MVB biogenesis and function. Specific Aim 4 proposes the characterization of DmVps28 as an example for such mutations acting early in the endocytic pathway. An important long-term goal of this work is to relate defects in different trafficking mutations to the symptoms in genetic diseases altering endocytic trafficking. To directly compare phenotypes in the Drosophila model system, Specific Aim 5 is directed towards the isolation of mutations in the Drosophila Hermansky-Pudlak syndrome-1 gene and the characterization of the resulting defects in endocytic trafficking.