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ROLE OF DROSOSPHIA PHOSPHOLIPASE D IN CELLULARIZATION

果蝇磷脂酶 D 在细胞化中的作用

基本信息

批准号：
6627272
负责人：
Michael A. Frohman
金额：
$ 20.94万
依托单位：
STATE UNIVERSITY NEW YORK STONY BROOK
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-01-01 至 2003-12-31
项目状态：
已结题

项目摘要

During the past several years, a superfamily of phosphatidylcholine-hydrolyzing Phospholipase D (PLD) genes conserved from prokaryotes to mammals has been described. Mammalian PLDs are activated by G-protein-coupled receptor and tyrosine kinase receptor signal transduction pathways. The biochemical step mediated by PLD and the second messenger generated by it are fairly well characterized, but functional roles for PLDs although thought to be important are not well defined except in yeast, where intriguing results have been generated. These results, together with less definitive mammalian studies, suggest that PLD promotes specialized types of membrane biogenesis as it relates to vesicular trafficking from the Golgi and plasma membrane. This proposal addresses potential roles of PLD in Drosophila melanogaster embryogenesis in cellularization, which is a specialized form of membrane biogenesis originating from the Golgi. We have mapped and cloned a PLD gene from Drosophila (denoted dPLD). Similar to mammalian PLD, dPLD appears to be regulated by Protein Kinase C-stimulated pathways. Unlike mammalian PLD but similar to yeast PLD, dPLD is not stimulated by the small G-protein ARF. dPLD-specific antisera reveal that it is expressed in germ cells and in the periphery of the embryo during cellularization. We propose 1) to define the mechanisms that regulate dPLD; 2) to characterize dPLD spatial expression and subcellular localization at high resolution in wild type embryos and embryos with defects in cellularization; 3) to generate and characterize loss-of-function dPLD alleles; 4) to examine the consequence of overexpression of wild-type and mutant PLD during cellularization and germ cell migration. Ultimately, we seek to understand what functional roles dPLD mediates, to model the more complex cell biological and physiological roles undertaken by the mammalian PLDs, and to bridge the growing knowledge concerning PLD mechanism of action in yeast and mammals using technological approaches unique to Drosophila.

在过去的几年中，磷脂酰胆碱水解磷脂酶D（PLD）基因的超家族从原核生物到哺乳动物的保守已被描述。哺乳动物PLD通过G蛋白偶联受体和酪氨酸激酶受体信号转导途径激活。 PLD介导的生化步骤和由它产生的第二信使是相当好的特点，但PLD的功能作用，虽然被认为是重要的是没有得到很好的定义，除了在酵母中，其中产生了有趣的结果。这些结果，连同不太确定的哺乳动物研究，表明PLD促进专门类型的膜生物合成，因为它涉及到从高尔基体和质膜的囊泡贩运。这一建议解决了潜在的作用PLD在果蝇胚胎细胞化，这是一个专门的形式的膜生物起源于高尔基体。我们已经定位并克隆了果蝇PLD基因（dPLD）。与哺乳动物PLD类似，dPLD似乎受蛋白激酶C刺激的途径调节。与哺乳动物PLD不同，但与酵母PLD相似，dPLD不受小G蛋白ARF的刺激。 dPLD特异性抗血清显示，它在生殖细胞和细胞化过程中胚胎的外周中表达。我们建议：1）定义调节dPLD的机制; 2）在野生型胚胎和细胞化缺陷的胚胎中以高分辨率表征dPLD空间表达和亚细胞定位; 3）产生和表征功能丧失的dPLD等位基因; 4）检查在细胞化和生殖细胞迁移过程中野生型和突变型PLD过表达的后果。最终，我们试图了解什么样的功能作用dPLD介导，模拟更复杂的细胞生物学和生理作用的哺乳动物PLD，并弥合不断增长的知识PLD的作用机制在酵母和哺乳动物中使用独特的技术方法果蝇。