UNIPARENTAL INHERITANCE OF CHLOROPLAST DNA

真核生物已经进化出几种策略来确保单亲 细胞器DNA的遗传。拟议工作的目标是 用实验方法阐明单亲遗传的机制 易驯服的生物体，单细胞藻类，莱茵衣藻。 叶绿体遗传受两个交配型等位基因控制 衣藻属、Mt+和Mt-。在单倍体细胞交配和减数分裂后， 后代只保留mt+叶绿体DNA。这一战略 衣藻用于维持单亲遗传似乎 涉及对合子mt+叶绿体DNA的选择性保护 在mt-和mt+叶绿体中都有活性的核酸酶。建议数 实验旨在分离该系统的关键组件，即 核酸酶和保护因子(S)。第一，有序列的菌株 将构建特定的叶绿体标签，以使mt- 可以对叶绿体DNA进行定量监测。第二，这些标签 这些菌株将被用于建立一种mt特异性的体外检测方法。 叶绿体DNA降解，以及mt+的特异性保护。这个化验结果会 用于纯化核酸酶和保护活性。最后， 核酸酶和保护活性有缺陷的突变株将 被分离和克隆。的生物化学和遗传学特征 这个系统将允许对潜在的分子机制进行剖析。 单亲继承。除了深入了解一个基本的 真核细胞器生物学的性质，这项工作还可能具有 对衰老和癌症的影响。的删除和换位 线粒体DNA和线粒体代谢的改变 与癌症和衰老过程有关。了解细胞如何 如Chalmydomonas消除了不受欢迎的细胞器基因组 提供有关细胞如何识别和调节的重要基本信息 它们细胞器DNA的命运。

Eukaryotes have evolved several strategies to ensure uniparental inheritance of organellar DNA. The goal of the proposed work is to elucidate the mechanism of uniparental inheritance using an experimentally tractable organism, the unicellular algae, Chlamydomonas reinhardtii. Chloroplast inheritance is controlled by the two mating type alleles in Chlamydomonas, mt+ and mt-. After haploid cells mate and undergo meiosis, the progeny retain only mt+ chloroplast DNA. The strategy that Chlamydomonas utilizes for maintaining uniparental inheritance appears to involve selective protection of mt+ chloroplast DNA from a zygotic nuclease that is active in both mt- and mt+ chloroplasts. The proposed experiments are designed to isolate the key components of this system, the nuclease and the protection factor(s). First, strains with sequence specific chloroplast tags will be constructed so that degradation of mt- chloroplast DNA can be quantitatively monitored. Second, these tagged strains will be used to develop an in vitro assay for mt- specific chloroplast DNA degradation, and mt+ specific protection. This assay will be used to purify the nuclease and protection activities. Finally, mutants that are defective for the nuclease and protection activities will be isolated and cloned. The biochemical and genetic characterization of this system will allow a dissection of the molecular mechanisms underlying uniparental inheritance. Besides gaining insight into a fundamental property of eukaryotic organellar biology, this work may also have implications for aging and cancer. Deletions and transpositions of mitochondrial DNA and alterations in mitochondrial metabolism are associated with cancer and the aging process. Understanding how a cell such as Chalmydomonas eliminates undesirable organellar genomes will provide important basic information on how cells recognize and regulate the fate of their organellar DNA.