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DEVELOPMENTAL GENETICS OF ASCIDIANS

海鞘的发育遗传学

基本信息

批准号：
12358012
负责人：
SATOH Noriyuki
金额：
$ 31.42万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2003
项目状态：
已结题

项目摘要

Ascidians provide an appealingly simple experimental system to investigate the function of developmentally relevant genes during chordate development. The ascidian tadpole is composed of only 〜2600 cells, which constitute a small set of larval organs including the epidermis, central nervous system, notochord and tail muscle. This configuration of the tadpole represents the basic chordate body plan. Embryogenesis of the best studied ascidians Ciona intestinalis and Ciona savignyi is rapid (about l8 hr), they are hermaphroditic, self-fertilization is feasible, and the entire life cycle takes less than 3 months, facilitating mutagenesis and genetic screens. The aim of the present study was to introduce forward genetic methods to isolate mutants in these ascidians.We first attempted mutagenesis with chemical mutagens. By treating sperm with the chemical mutagen ENU, several mutant lines were established. For example, chongmague affects notochord formation and zebulon affects head formation … More in Ciona intestinalis embryos.The identification of the affected genes is not simple when mutations are caused by single nucleotide substitutions. A strategy for identifying mutated genes involves insertional mutagenesis with the aid of transposable elements. However, previous attempts at insertional mutagenesis in marine invertebrates were not successful. We found that a Tc1/mariner family transposon Minos, originally isolated from Drosophila hydei, can function as a transposable element in Ciona. The Ciona intestinalis genome is AT-rich and Minos targets TA dinucleotides. This may explain why Minos is active in Ciona embryos. By careful examination of excision, deletion and transposable activity, we have established a number of enhancer trap lines in Ciona intestinalis. This method might represent a significant breakthrough for elucidating gene function in Ciona. At present, we are isolating mutants with deficiencies of adult organ formation.Thus, the present study is the first report of possible insertional mutagenesis in marine animals. Less

海鞘提供了一个吸引人的简单实验系统来研究发育相关基因在脊索发育过程中的功能。海鞘蝌蚪仅由~2600个细胞组成，这些细胞构成了包括表皮、中枢神经系统、脊索和尾肌在内的一小套幼虫器官。蝌蚪的这种构型代表了基本的弦体平面。研究得最好的海鞘类海鞘的胚胎发生速度很快(约18小时)，它们是两性的，自交是可行的，整个生命周期只需不到3个月，有利于突变和遗传筛选。本研究的目的是介绍正向遗传学方法来分离这些海鞘中的突变体。我们首先尝试用化学诱变剂进行诱变。用化学诱变剂ENU处理精子，建立了几个突变株系。例如，冲马术影响脊索形成，而泽布隆影响头部形成…。当单核苷酸替换引起突变时，识别受影响的基因并不简单。一种识别突变基因的策略包括借助转座元件进行插入突变。然而，之前在海洋无脊椎动物中进行插入突变的尝试都没有成功。我们发现Tc1/Mariner家族转座子Minos最初是从果蝇中分离出来的，在Ciona中可以作为转座子元件发挥作用。海鞘基因组富含AT，而Minos则以TA二核苷酸为靶标。这可能解释了为什么Minos在Ciona胚胎中是活跃的。通过对基因的切除、缺失和转座活性的仔细研究，我们建立了一系列的肠上皮细胞增强子捕捉系。该方法可能是对芝麻属基因功能研究的重大突破。目前，我们正在分离成年器官形成缺陷的突变体。因此，本研究是海洋动物中可能的插入突变的第一个报告。较少