Evolution of Dosage Compensation for Amylase in Drosophila

果蝇淀粉酶剂量补偿的演变

基本信息

  • 批准号:
    8806510
  • 负责人:
  • 金额:
    $ 21.32万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
    Continuing Grant
  • 财政年份:
    1988
  • 资助国家:
    美国
  • 起止时间:
    1988-08-01 至 1992-07-31
  • 项目状态:
    已结题

项目摘要

Sex-linked dosage compensation is a mechanism of gene regulation that results in the level of X-linked gene product being equal in both sexes, although males have a single active gene copy for the product while females have two. In Drosophila miranda, the structural gene for alpha-amylase, Amy, is sex- linked and located in a relatively "new" X chromosome, X2, which evolved in response to an ancestral translocation of one arm of an autosome to the Y chromosome. The unexpressed homolog of Amy is in the "neo-Y" chromosome of males. Male and female larvae express equivalent levels of amylase activity, suggesting full dosage compensation at this developmental stage. By contrast, the compensatory mechanism appears to have been, at best, only partially evolved in adults. Since Amy is not sex-linked in very closely related Drosophila species - pseudoobscura and persimilis, this group of three sibling species provides an opportunity to study evolution of a eukaryotic gene regulatory mechanism. The ultimate goal is to understand the molecular basis of dosage compensation for amylase in D. miranda and how it may have evolved through DNA sequence changes. Doane and Norman have cloned and partially characterized the Amy region of miranda and isolated a clone that contains the functional Amy gene from its X2 chromosome. They also have begun to characterize dosage compensation of Amy expression at the level of amylase messenger RNA. The aims of this proposal are to: (1) complete the amylase RNA studies and assay dosage compensation at the RNA level in male midguts during development; (2) clone and characterize the autosomal Amy region of a selected strain of D. persimilis; (3) sequence and compare the functional Amy gene and a putative Amy pseudogene from miranda and compare the former sequence with the functional Amy sequence of persimilis; (4) attempt to clone and characterize Amy-homologous sequences from the neo-Y chromosome of miranda; (5) establish a radiation-induced Amy null (no amylase) strain of miranda to provide hosts for Amy gene transformation experiments; and (6) delineate the regulatory sequences that are important in dosage compensation of Amy expression in males of miranda through transformation experiments coupled with the deletion or alteration of specific DNA sequences adjacent to the amylase coding region. Transformation experiments, intra- and interspecific, will include both transient expression assays ( somatic transformations) and P. element-induced germline transformations, both of which entail the injection of DNA sequences to be tested in Amy null host embryos..Z. The significance of this basic research lies in the limited understanding of gene regulatory mechanisms in higher animals. This is especially true of mechanisms like Drosophila dosage compensation which augments activity of an already transcriptionally active gene. The opportunity provided by the amylase system is unique in that the compensatory mechanism is not yet fully evolved, so an intermediate stage in DNA sequence reorganization leading to it may be analyzed. Information about this should add significantly to our understanding of how gene regulatory mechanisms may evolve and how important they may be to the process of evolution. Such knowledge would be of potential value in the application of genetic engineering techniques to species in which it would be useful to alter the expression of a given gene for beneficial or economic reasons, or for experimental purposes in population studies.
性别连锁剂量补偿是一种基因调控机制,导致两性中x连锁基因产物的水平相等,尽管雄性有一个活性基因副本,而雌性有两个。在米兰达果蝇中,α -淀粉酶的结构基因Amy是性别连锁的,位于一个相对“新的”X染色体X2上,这是由于祖先将常染色体的一条臂易位到Y染色体上而进化而来的。Amy未表达的同系物位于男性的“新y”染色体中。雄性和雌性幼虫表现出相当水平的淀粉酶活性,表明在这个发育阶段有充分的剂量补偿。相比之下,补偿机制似乎在成人中充其量只是部分进化。由于Amy在亲缘关系非常密切的果蝇物种(拟眼果蝇和persimilis)中没有性别联系,这组三个兄弟物种为研究真核基因调控机制的进化提供了机会。最终目的是了解D. miranda中淀粉酶剂量补偿的分子基础,以及它如何通过DNA序列变化而进化。多恩和诺曼已经克隆了米兰达的艾米区,并对其进行了部分表征,并从其X2染色体中分离出了一个含有功能性艾米基因的克隆体。他们也开始在淀粉酶信使RNA水平上表征Amy表达的剂量补偿。本研究的目的是:(1)完成雄性中肠发育过程中淀粉酶RNA的研究和测定RNA水平上的剂量补偿;(2)克隆一株persimilis的常染色体Amy区并对其进行鉴定;(3)测序并比较功能性Amy基因和来自miranda的推定Amy假基因,并将前者序列与persimilis的功能性Amy序列进行比较;(4)尝试从miranda的neo-Y染色体中克隆并鉴定amy同源序列;(5)建立辐射诱导的无淀粉酶miranda菌株,为Amy基因转化实验提供宿主;(6)通过结合淀粉酶编码区附近特定DNA序列的缺失或改变的转化实验,描述了在miranda雄性中Amy表达剂量补偿中重要的调控序列。转化实验,种内和种间,将包括瞬时表达分析(体细胞转化)和p元素诱导的种系转化,这两种转化都需要注射DNA序列,以便在Amy null宿主胚胎中进行测试。这项基础研究的意义在于对高等动物基因调控机制的认识有限。这在果蝇的剂量补偿机制中尤其正确,它增加了已经具有转录活性的基因的活性。淀粉酶系统提供的机会是独一无二的,因为代偿机制尚未完全进化,因此可以分析导致它的DNA序列重组的中间阶段。这方面的信息将大大增加我们对基因调控机制如何进化的理解,以及它们对进化过程的重要性。这些知识在将基因工程技术应用于物种方面具有潜在价值,因为出于有益的或经济的原因,或为了人口研究中的实验目的,改变某一特定基因的表达将是有用的。

项目成果

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Winifred Doane其他文献

Winifred Doane的其他文献

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{{ truncateString('Winifred Doane', 18)}}的其他基金

REU: Evolution of Dosage Compensation for Amylase in Drosophilia miranda
REU:米兰达果蝇淀粉酶剂量补偿的演变
  • 批准号:
    8613579
  • 财政年份:
    1987
  • 资助金额:
    $ 21.32万
  • 项目类别:
    Standard Grant
Workshop on Drosophila Alpha-Amylases April 10, 1986, Asilomar, California
果蝇 α-淀粉酶研讨会 1986 年 4 月 10 日,加利福尼亚州阿西洛玛
  • 批准号:
    8600824
  • 财政年份:
    1986
  • 资助金额:
    $ 21.32万
  • 项目类别:
    Standard Grant

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剂量补偿的演变 - 使用具有独立进化 XX/XY 和 ZZ/ZW 染色体的海龟进行的实证测试
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CAREER: Evolution of allosomes and dosage compensation in terrestrial isopods
职业:陆地等足动物异体的进化和剂量补偿
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The evolution of dosage compensation in Drosophila
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